Proteopedia - User contributions [en]

Proteopedia:Page of the Year Entrants

2009-12-19T18:55:32Z

Maitreyee Mukherjee:

Please list your entry in [[Proteopedia:Page of the Year Competition| Proteopedia's Page of the Year Competition]] here. You can enter as many pages as you like.
Add your entry to the list by adding a row in the format shown below, with a link to your user page followed by 2 hyphens and then a link to the page you're entering in the competition.

----

==Entrants==

Entry number 1 is just an example, Eran cannot participate in the competition.

# [[User:Eran Hodis]] -- [[User:Eran_Hodis/Acetylcholinesterase]]
# [[User:Ramiro Barrantes]] -- [[1tdh]]
# [[User:Sara Toftegaard Petersen]] [[User:Mathilde Thomsen]] [[User:Mette Trauelsen]] -- [[Nitric oxide synthase]]
# [[User:Wayne Decatur]] -- [[Ribosome]] (see history for original article as of October 15, 2009)
# [[User:Wayne Decatur]] -- [[User:Wayne Decatur/Plant Viral Protein p19 Suppression of RNA Silencing]]
# [[User:Wayne Decatur]] -- [[User:Wayne Decatur/Suppression of RNA Silencing by Viruses]]
# [[User:Wayne Decatur]] -- [[User:Wayne Decatur/Tomato aspermy virus protein 2b Suppression of RNA Silencing]]
# [[User:Tzviya Zeev-Ben-Mordehai]] -- [[Intrinsically Disordered Protein]] (see history for original article as of October 19, 2009)
# [[User:Tilman Schirmer]] -- [[C-di-GMP_signaling]] (Note that there are several sub-pages)
# [[User:Joseph Lipsick]] -- [[SRC]]
#[[User:Maitreyee Mukherjee]]--[[http://www.proteopedia.org/wiki/index.php/PHB_synthase_in_Rhodobacter_sphaeroides]]

Proteopedia:Page of the Year Entrants

2009-12-19T18:54:50Z

Maitreyee Mukherjee:

Please list your entry in [[Proteopedia:Page of the Year Competition| Proteopedia's Page of the Year Competition]] here. You can enter as many pages as you like.
#[[User:Maitreyee Mukherjee]]--[[http://www.proteopedia.org/wiki/index.php/PHB_synthase_in_Rhodobacter_sphaeroides]]
Add your entry to the list by adding a row in the format shown below, with a link to your user page followed by 2 hyphens and then a link to the page you're entering in the competition.

----

==Entrants==

Entry number 1 is just an example, Eran cannot participate in the competition.

# [[User:Eran Hodis]] -- [[User:Eran_Hodis/Acetylcholinesterase]]
# [[User:Ramiro Barrantes]] -- [[1tdh]]
# [[User:Sara Toftegaard Petersen]] [[User:Mathilde Thomsen]] [[User:Mette Trauelsen]] -- [[Nitric oxide synthase]]
# [[User:Wayne Decatur]] -- [[Ribosome]] (see history for original article as of October 15, 2009)
# [[User:Wayne Decatur]] -- [[User:Wayne Decatur/Plant Viral Protein p19 Suppression of RNA Silencing]]
# [[User:Wayne Decatur]] -- [[User:Wayne Decatur/Suppression of RNA Silencing by Viruses]]
# [[User:Wayne Decatur]] -- [[User:Wayne Decatur/Tomato aspermy virus protein 2b Suppression of RNA Silencing]]
# [[User:Tzviya Zeev-Ben-Mordehai]] -- [[Intrinsically Disordered Protein]] (see history for original article as of October 19, 2009)
# [[User:Tilman Schirmer]] -- [[C-di-GMP_signaling]] (Note that there are several sub-pages)
# [[User:Joseph Lipsick]] -- [[SRC]]

PHB synthase in Rhodobacter sphaeroides

2009-05-04T05:07:23Z

Maitreyee Mukherjee: New page: <applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain in Rhodobacter sphaeroides,amino acids 286 to 547' /> =Introduct...

User:Maitreyee Mukherjee/Sandbox 1

2009-05-04T05:03:51Z

Maitreyee Mukherjee:

User:Maitreyee Mukherjee/Sandbox 1

2009-05-04T05:00:01Z

Maitreyee Mukherjee:

User:Maitreyee Mukherjee/Sandbox 1

2009-05-03T19:52:44Z

Maitreyee Mukherjee:

User:Maitreyee Mukherjee/Sandbox 1

2009-04-30T00:36:20Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain in Rhodobacter sphaeroides,amino acids 286 to 547' />

=Introduction=

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria<ref>PMID: 14999403</ref> which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products<ref>PMID:10416645</ref>. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate <ref> Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of Rhodobacter sphaeroides. Biotechnology Letters. 15:709–714.</ref>.''Rhodobacter sphaeroides'' possesses the Class I phylogenetic group of PHA synthases<ref>PMID:14620841</ref>. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms<ref>PMID:12954080</ref>.

=Function=

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria.

=Properties=

Clicking on the green links will show different forms of the structure. A description of what the colors mean are given below the green links.

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/4'>Ball and Stick model in N>C rainbow color</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/5'>Secondary Structure</scene>

{{Template:ColorKey_Helix}},
{{Template:ColorKey_Strand}},
{{Template:ColorKey_Turn}}.

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

=Current Research: Knowledge of PHB synthase in ''R.sphaeroides'' as well as in other organisms=

1. In ''R.sphaeroides'' FJ1, ''phbC'' has been found to be present on one of two gene locus identified to be involved in the production of PHB. The gene ''phbC'' is present on the same locus along with the other genes responsible for PHB syn thesis such as ''phbZ'', ''phbP'' and ''phbR''. <ref>PMID: 16440119</ref>

2. In a paper by Fales et.al. in 2001, the investigators talk about a possible connection between PHB synthase activity and ''hemA'' transcription in ''R.sphaeroides''. No further investigations answering this have been published after this untill now. The investigators also present a schematic diagram of a proposed dual pathway of PHB synthesis in this organism.<ref>PMID: 11160087</ref>

3. In ''Rhodobacter capsulatus'', ''phaC'' was found to have a constitutive expression in presence or absence of the induction of PHA production, indicating that the control of expression of this gene occurs at the post-translational level in this organism. <ref>PMID: 9251189</ref>

4. In ''Paracoccus denitrificans'', an organism having a close similarity in genome to ''R.sphaeroides'', the ''phaC'' transcription has been found to be reduced under nitrogen deficient condition, although the mechanism of regulation of this gene is not very clear yet..<ref>PMID: 16233588</ref>. in the cited paper, the investigators have included a proposed model for regulation of PHB synthesis in this organism.

5. One group of investigators suggest a role of the genes involved in the nitrogen-related phosphotransferase system (PTS)in PHB accumulation in the organism ''Azotobacter vinelandii''. This group of researchers report that one of the proteins of PTS affect the transcription of the ''phbBAC'' operon(consisting of the ''phbA'', ''phbB'' and ''phbC'' genes) in this organism. Mechanism of this regulation is still unclear.<ref>PMID:17878711</ref>

=Probable Reseach:Challenges towards our Understanding=

1. What is the molecular basis of regulation of PHB synthesis in this organism and what is the role of this protein in it?

2. How does environmental factors such as presence or absence of oxygen or nitrogen affect the activity of this protein in ''R.sphaeroides''?

3. What are the activators or repressor factors, if any, responsible for production of this protein in this organism?

4. Is there a possible role of the nitrogen-related Phosphotransferase system in the regulation of expression of PHB synthase in ''R.sphaeroides''?

=How was the animated image generated?=

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

=How was the JMOL image generated?=

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

=Links to images on this page=

http://bmm.cancerresearchuk.org/~3djigsaw/dom_fish/display2_maestro.cgi?output/fabdae5e, JMOL Image.

http://polyview.cchmc.org/cgi-bin/get_image_3d.cgi?IMG=12407603974718, animated image.

=References=

<references/>

PAGE CREATED BY MAITREYEE MUKHERJEE, email:maitreyee25@gmail.com

ACKNOWLEDGEMENT: Jill Zeilstra Ryalls, Professor, Bowling Green State University.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-30T00:33:44Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain in Rhodobacter sphaeroides,amino acids 286 to 547' />

=Introduction=

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria<ref>PMID: 14999403</ref> which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products<ref>PMID:10416645</ref>. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate <ref> Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of Rhodobacter sphaeroides. Biotechnology Letters. 15:709–714.</ref>.''Rhodobacter sphaeroides'' possesses the Class I phylogenetic group of PHA synthases<ref>PMID:14620841</ref>. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms<ref>PMID:12954080</ref>.

=Function=

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria.

=Properties=

Clicking on the green links will show different forms of the structure. A description of what the colors mean are given below the green links.

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/4'>Ball and Stick model in N>C rainbow color</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/5'>Secondary Structure</scene>

{{Template:ColorKey_Helix}},
{{Template:ColorKey_Strand}},
{{Template:ColorKey_Turn}}.

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

=Current Research: Knowledge of PHB synthase in ''R.sphaeroides'' as well as in other organisms=

1. In ''R.sphaeroides'' FJ1, ''phbC'' has been found to be present on one of two gene locus identified to be involved in the production of PHB. The gene ''phbC'' is present on the same locus along with the other genes responsible for PHB syn thesis such as ''phbZ'', ''phbP'' and ''phbR''. <ref>PMID: 16440119</ref>

2. In a paper by Fales et.al. in 2001, the investigators talk about a possible connection between PHB synthase activity and ''hemA'' transcription in ''R.sphaeroides''. No further investigations answering this have been published after this untill now. The investigators also present a schematic diagram of a proposed dual pathway of PHB synthesis in this organism.<ref>PMID: 11160087</ref>

3. In ''Rhodobacter capsulatus'', ''phaC'' was found to have a constitutive expression in presence or absence of the induction of PHA production, indicating that the control of expression of this gene occurs at the post-translational level in this organism. <ref>PMID: 9251189</ref>

4. In ''Paracoccus denitrificans'', an organism having a close similarity in genome to ''R.sphaeroides'', the ''phaC'' transcription has been found to be reduced under nitrogen deficient condition, although the mechanism of regulation of this gene is not very clear yet..<ref>PMID: 16233588</ref>. in the cited paper, the investigators have included a proposed model for regulation of PHB synthesis in this organism.

5. One group of investigators suggest a role of the genes involved in the nitrogen-related phosphotransferase system (PTS)in PHB accumulation in the organism ''Azotobacter vinelandii''. This group of researchers report that one of the proteins of PTS affect the transcription of the ''phbBAC'' operon(consisting of the ''phbA'', ''phbB'' and ''phbC'' genes) in this organism. Mechanism of this regulation is still unclear.<ref>PMID:17878711</ref>

=Probable Reseach:For Better Understanding of PHB synthase:Challenges towards our Understanding=

1. What is the molecular basis of regulation of PHB synthesis in this organism and what is the role of this protein in it?

2. How does environmental factors such as presence or absence of oxygen or nitrogen affect the activity of this protein in ''R.sphaeroides''?

3. What are the activators or repressor factors, if any, responsible for production of this protein in this organism?

4. Is there a possible role of the nitrogen-related Phosphotransferase system in the regulation of expression of PHB synthase in ''R.sphaeroides''?

=How was the animated image generated?=

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

=How was the JMOL image generated?=

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

=Links to images on this page=

http://bmm.cancerresearchuk.org/~3djigsaw/dom_fish/display2_maestro.cgi?output/fabdae5e, JMOL Image.

http://polyview.cchmc.org/cgi-bin/get_image_3d.cgi?IMG=12407603974718, animated image.

=References=

<references/>

PAGE CREATED BY MAITREYEE MUKHERJEE, email:maitreyee25@gmail.com

ACKNOWLEDGEMENT: Jill Zeilstra Ryalls, Professor, Bowling Green State University.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-30T00:32:41Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain in Rhodobacter sphaeroides,amino acids 286 to 547' />

=Introduction=

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria<ref>PMID: 14999403</ref> which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products<ref>PMID:10416645</ref>. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate <ref> Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of Rhodobacter sphaeroides. Biotechnology Letters. 15:709–714.</ref>.''Rhodobacter sphaeroides'' possesses the Class I phylogenetic group of PHA synthases<ref>PMID:14620841</ref>. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms<ref>PMID:12954080</ref>.

=Function=

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria.

=Properties=

Clicking on the green links will show different forms of the structure. A description of what the colors mean are given below the green links.

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/4'>Ball and Stick model in N>C rainbow color</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/5'>Secondary Structure</scene>

{{Template:ColorKey_Helix}},
{{Template:ColorKey_Strand}},
{{Template:ColorKey_Turn}}.

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

=Current research: Knowledge of PHB synthase in ''R.sphaeroides'' as well as in other organisms=

1. In ''R.sphaeroides'' FJ1, ''phbC'' has been found to be present on one of two gene locus identified to be involved in the production of PHB. The gene ''phbC'' is present on the same locus along with the other genes responsible for PHB syn thesis such as ''phbZ'', ''phbP'' and ''phbR''. <ref>PMID: 16440119</ref>

2. In a paper by Fales et.al. in 2001, the investigators talk about a possible connection between PHB synthase activity and ''hemA'' transcription in ''R.sphaeroides''. No further investigations answering this have been published after this untill now. The investigators also present a schematic diagram of a proposed dual pathway of PHB synthesis in this organism.<ref>PMID: 11160087</ref>

3. In ''Rhodobacter capsulatus'', ''phaC'' was found to have a constitutive expression in presence or absence of the induction of PHA production, indicating that the control of expression of this gene occurs at the post-translational level in this organism. <ref>PMID: 9251189</ref>

4. In ''Paracoccus denitrificans'', an organism having a close similarity in genome to ''R.sphaeroides'', the ''phaC'' transcription has been found to be reduced under nitrogen deficient condition, although the mechanism of regulation of this gene is not very clear yet..<ref>PMID: 16233588</ref>. in the cited paper, the investigators have included a proposed model for regulation of PHB synthesis in this organism.

5. One group of investigators suggest a role of the genes involved in the nitrogen-related phosphotransferase system (PTS)in PHB accumulation in the organism ''Azotobacter vinelandii''. This group of researchers report that one of the proteins of PTS affect the transcription of the ''phbBAC'' operon(consisting of the ''phbA'', ''phbB'' and ''phbC'' genes) in this organism. Mechanism of this regulation is still unclear.<ref>PMID:17878711</ref>

=Probable Reseach:For Better Understanding of PHB synthase:Challenges towards our Understanding=

1. What is the molecular basis of regulation of PHB synthesis in this organism and what is the role of this protein in it?

2. How does environmental factors such as presence or absence of oxygen or nitrogen affect the activity of this protein in ''R.sphaeroides''?

3. What are the activators or repressor factors, if any, responsible for production of this protein in this organism?

4. Is there a possible role of the nitrogen-related Phosphotransferase system in the regulation of expression of PHB synthase in ''R.sphaeroides''?

=How was the animated image generated?=

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

=How was the JMOL image generated?=

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

=Links to images on this page=

http://bmm.cancerresearchuk.org/~3djigsaw/dom_fish/display2_maestro.cgi?output/fabdae5e, JMOL Image.

http://polyview.cchmc.org/cgi-bin/get_image_3d.cgi?IMG=12407603974718, animated image.

=References=

<references/>

PAGE CREATED BY MAITREYEE MUKHERJEE, email:maitreyee25@gmail.com

ACKNOWLEDGEMENT: Jill Zeilstra Ryalls, Professor, Bowling Green State University.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-30T00:29:28Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain in Rhodobacter sphaeroides,amino acids 286 to 547' />

=Introduction=

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria<ref>PMID: 14999403</ref> which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products<ref>PMID:10416645</ref>. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate <ref> Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of Rhodobacter sphaeroides. Biotechnology Letters. 15:709–714.</ref>.''Rhodobacter sphaeroides'' possesses the Class I phylogenetic group of PHA synthases<ref>PMID:14620841</ref>. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms<ref>PMID:12954080</ref>.

=Function=

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria.

=Properties=

Clicking on the green links will show different forms of the structure. A description of what the colors mean are given below the green links.

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/4'>Ball and Stick model in N>C rainbow color</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/5'>Secondary Structure</scene>

{{Template:ColorKey_Helix}},
{{Template:ColorKey_Strand}},
{{Template:ColorKey_Turn}}.

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

=Current research: Knowledge of PHB synthase in ''R.sphaeroides'' as well as in other organisms=

1. In ''R.sphaeroides'' FJ1, ''phbC'' has been found to be present on one of two gene locus identified to be involved in the production of PHB. The gene ''phbC'' is present on the same locus along with the other genes responsible for PHB syn thesis such as ''phbZ'', ''phbP'' and ''phbR''. <ref>PMID: 16440119</ref>

2. In a paper by Fales et.al. in 2001, the investigators talk about a possible connection between PHB synthase activity and ''hemA'' transcription in ''R.sphaeroides''. No further investigations answering this have been published after this untill now. The investigators also present a schematic diagram of a proposed dual pathway of PHB synthesis in this organism.<ref>PMID: 11160087</ref>

3. In ''Rhodobacter capsulatus'', ''phaC'' was found to have a constitutive expression in presence or absence of the induction of PHA production, indicating that the control of expression of this gene occurs at the post-translational level in this organism. <ref>PMID: 9251189</ref>

4. In ''Paracoccus denitrificans'', an organism having a close similarity in genome to ''R.sphaeroides'', the ''phaC'' transcription has been found to be reduced under nitrogen deficient condition, although the mechanism of regulation of this gene is not very clear yet..<ref>PMID: 16233588</ref>. in the cited paper, the investigators have included a proposed model for regulation of PHB synthesis in this organism.

5. One group of investigators suggest a role of the genes involved in the nitrogen-related phosphotransferase system (PTS)in PHB accumulation in the organism ''Azotobacter vinelandii''. This group of researchers report that one of the proteins of PTS affect the transcription of the ''phbBAC'' operon(consisting of the ''phbA'', ''phbB'' and ''phbC'' genes) in this organism. Mechanism of this regulation is still unclear.<ref>PMID:17878711</ref>

=Probable Reseach:For Better Understanding of PHB synthase:Chanllenges towards our Understanding=

1. What is the molecular basis of regulation of PHB synthesis in this organism and what is the role of this protein in it?

2. How does environmental factors such as presence or absence of oxygen or nitrogen relate to the activity of this protein in ''R.sphaeroides''?

3. What are the activators or repressor factors, if any, responsible for production of this protein in this organism?

4. Is there a possible role of the nitrogen-related Phosphotransferase system in the regulation of expression of PHB synthase in ''R.sphaeroides''?

=How was the animated image generated?=

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

=How was the JMOL image generated?=

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

=Links to images on this page=

http://bmm.cancerresearchuk.org/~3djigsaw/dom_fish/display2_maestro.cgi?output/fabdae5e, JMOL Image.

http://polyview.cchmc.org/cgi-bin/get_image_3d.cgi?IMG=12407603974718, animated image.

=References=

<references/>

PAGE CREATED BY MAITREYEE MUKHERJEE, email:maitreyee25@gmail.com

ACKNOWLEDGEMENT: Jill Zeilstra Ryalls, Professor, Bowling Green State University.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-30T00:24:52Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain in Rhodobacter sphaeroides,amino acids 286 to 547' />

=Introduction=

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria<ref>PMID: 14999403</ref> which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products<ref>PMID:10416645</ref>. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate <ref> Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of Rhodobacter sphaeroides. Biotechnology Letters. 15:709–714.</ref>.''Rhodobacter sphaeroides'' possesses the Class I phylogenetic group of PHA synthases<ref>PMID:14620841</ref>. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms<ref>PMID:12954080</ref>.

=Function=

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria.

=Properties=

Clicking on the green links will show different forms of the structure. A description of what the colors mean are given below the green links.

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/4'>Ball and Stick model in N>C rainbow color</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/5'>Secondary Structure</scene>

{{Template:ColorKey_Helix}},
{{Template:ColorKey_Strand}},
{{Template:ColorKey_Turn}}.

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

=Current research: Knowledge of PHB synthase in ''R.sphaeroides'' as well as in other organisms=

1. In ''R.sphaeroides'' FJ1, ''phbC'' has been found to be present on one of two gene locus identified to be involved in the production of PHB. The gene ''phbC'' is present on the same locus along with the other genes responsible for PHB syn thesis such as ''phbZ'', ''phbP'' and ''phbR''. <ref>PMID: 16440119</ref>

2. In a paper by Fales et.al. in 2001, the investigators talk about a possible connection between PHB synthase activity and ''hemA'' transcription in ''R.sphaeroides''. No further investigations answering this have been published after this untill now. The investigators also present a schematic diagram of a proposed dual pathway of PHB synthesis in this organism.<ref>PMID: 11160087</ref>

3. In ''Rhodobacter capsulatus'', ''phaC'' was found to have a constitutive expression in presence or absence of the induction of PHA production, indicating that the control of expression of this gene occurs at the post-translational level in this organism. <ref>PMID: 9251189</ref>

4. In ''Paracoccus denitrificans'', an organism having a close similarity in genome to ''R.sphaeroides'', the ''phaC'' transcription has been found to be reduced under nitrogen deficient condition, although the mechanism of regulation of this gene is not very clear yet..<ref>PMID: 16233588</ref>. in the cited paper, the investigators have included a proposed model for regulation of PHB synthesis in this organism.

5. One group of investigators suggest a role of the genes involved in the nitrogen-related phosphotransferase system (PTS)in PHB accumulation in the organism ''Azotobacter vinelandii''. This group of researchers report that one of the proteins of PTS affect the transcription of the ''phbBAC'' operon(consisting of the ''phbA'', ''phbB'' and ''phbC'' genes) in this organism. Mechanism of this regulation is still unclear.<ref>PMID:17878711</ref>

=Probable Reseach:For Better Understanding of PHB synthase:Chanllenges towards our Understanding=

1. What is the molecular basis of regulation of PHB synthesis in this organism and what is the role of this protein in it?

2. How does environmental factors such as presence or absence of oxygen or nitrogen relate to the activity of this protein in ''R.sphaeroides''?

3. What are the activators or repressor factors, if any, responsible for production of this protein in this organism?

4. Is there a possible role of the nitrogen-related Phosphotransferase system in the regulation of expression of PHB synthase in ''R.sphaeroides''?

=How was the animated image generated?=

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

=How was the JMOL image generated?=

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

=Links to images on this page=

http://bmm.cancerresearchuk.org/~3djigsaw/dom_fish/display2_maestro.cgi?output/fabdae5e, JMOL Image.

http://polyview.cchmc.org/cgi-bin/get_image_3d.cgi?IMG=12407603974718, animated image.

=References=

<references/>

PAGE CREATED BY MAITREYEE MUKHERJEE, email:maitreyee25@gmail.com

ACKNOWLEDGEMENT: Jill Zeilstra Ryalls, Professor, Bowling Green State University.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-30T00:22:34Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain in Rhodobacter sphaeroides,amino acids 286 to 547' />

=Introduction=

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria<ref>PMID: 14999403</ref> which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products<ref>PMID:10416645</ref>. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate <ref> Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of Rhodobacter sphaeroides. Biotechnology Letters. 15:709–714.</ref>.''Rhodobacter sphaeroides'' possesses the Class I phylogenetic group of PHA synthases<ref>PMID:14620841</ref>. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms<ref>PMID:12954080</ref>.

=Function=

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria.

=Properties=

Clicking on the green links will show different forms of the structure. description of what the colors mean are given below the green links.

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/4'>Ball and Stick model in N>C rainbow color</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/5'>Secondary Structure</scene>

{{Template:ColorKey_Helix}},
{{Template:ColorKey_Strand}},
{{Template:ColorKey_Turn}}.

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

=Current research: Knowledge of PHB synthase in ''R.sphaeroides'' as well as in other organisms=

1. In ''R.sphaeroides'' FJ1, ''phbC'' has been found to be present on one of two gene locus identified to be involved in the production of PHB. The gene ''phbC'' is present on the same locus along with the other genes responsible for PHB syn thesis such as ''phbZ'', ''phbP'' and ''phbR''. <ref>PMID: 16440119</ref>

2. In a paper by Fales et.al. in 2001, the investigators talk about a possible connection between PHB synthase activity and ''hemA'' transcription in ''R.sphaeroides''. No further investigations answering this have been published after this untill now. The investigators also present a schematic diagram of a proposed dual pathway of PHB synthesis in this organism.<ref>PMID: 11160087</ref>

3. In ''Rhodobacter capsulatus'', ''phaC'' was found to have a constitutive expression in presence or absence of the induction of PHA production, indicating that the control of expression of this gene occurs at the post-translational level in this organism. <ref>PMID: 9251189</ref>

4. In ''Paracoccus denitrificans'', an organism having a close similarity in genome to ''R.sphaeroides'', the ''phaC'' transcription has been found to be reduced under nitrogen deficient condition, although the mechanism of regulation of this gene is not very clear yet..<ref>PMID: 16233588</ref>. in the cited paper, the investigators have included a proposed model for regulation of PHB synthesis in this organism.

5. One group of investigators suggest a role of the genes involved in the nitrogen-related phosphotransferase system (PTS)in PHB accumulation in the organism ''Azotobacter vinelandii''. This group of researchers report that one of the proteins of PTS affect the transcription of the ''phbBAC'' operon(consisting of the ''phbA'', ''phbB'' and ''phbC'' genes) in this organism. Mechanism of this regulation is still unclear.<ref>PMID:17878711</ref>

=Probable Reseach:For Better Understanding of PHB synthase:Chanllenges towards our Understanding=

1. What is the molecular basis of regulation of PHB synthesis in this organism and what is the role of this protein in it?

2. How does environmental factors such as presence or absence of oxygen or nitrogen relate to the activity of this protein in ''R.sphaeroides''?

3. What are the activators or repressor factors, if any, responsible for production of this protein in this organism?

4. Is there a possible role of the nitrogen-related Phosphotransferase system in the regulation of expression of PHB synthase in ''R.sphaeroides''?

=How was the animated image generated?=

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

=How was the JMOL image generated?=

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

=Links to images on this page=

http://bmm.cancerresearchuk.org/~3djigsaw/dom_fish/display2_maestro.cgi?output/fabdae5e, JMOL Image.

http://polyview.cchmc.org/cgi-bin/get_image_3d.cgi?IMG=12407603974718, animated image.

=References=

<references/>

PAGE CREATED BY MAITREYEE MUKHERJEE, email:maitreyee25@gmail.com

ACKNOWLEDGEMENT: Jill Zeilstra Ryalls, Professor, Bowling Green State University.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-27T15:57:11Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain in Rhodobacter sphaeroides,amino acids 286 to 547' />

=Introduction=

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria<ref>PMID: 14999403</ref> which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products<ref>PMID:10416645</ref>. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate <ref> Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of Rhodobacter sphaeroides. Biotechnology Letters. 15:709–714.</ref>.''Rhodobacter sphaeroides'' possesses the Class I phylogenetic group of PHA synthases<ref>PMID:14620841</ref>. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms<ref>PMID:12954080</ref>.

=Function=

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria.

'''PROPERTIES:'''

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/4'>ball and stick model</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/5'>secondary structure</scene>

{{Template:ColorKey_Helix}},
{{Template:ColorKey_Strand}},
{{Template:ColorKey_Turn}}.

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''CURRENT RESEARCH: KNOWLEDGE OF THIS PROTEIN IN ''R.sphaeroides'' AS WELL AS IN OTHER ORGANISMS:'''

1. In ''R.sphaeroides'' FJ1, ''phbC'' has been found to be present on one of two gene locus identified to be involved in the production of PHB. The gene ''phbC'' is present on the same locus along with the other genes responsible for PHB syn thesis such as ''phbZ'', ''phbP'' and ''phbR''. <ref>PMID: 16440119</ref>

2. In a paper by Fales et.al. in 2001, the investigators talk about a possible connection between PHB synthase activity and ''hemA'' transcription in ''R.sphaeroides''. No further investigations answering this have been published after this untill now. The investigators also present a schematic diagram of a proposed dual pathway of PHB synthesis in this organism.<ref>PMID: 11160087</ref>

3. In ''Rhodobacter capsulatus'', ''phaC'' was found to have a constitutive expression in presence or absence of the induction of PHA production, indicating that the control of expression of this gene occurs at the post-translational level in this organism. <ref>PMID: 9251189</ref>

4. In ''Paracoccus denitrificans'', an organism having a close similarity in genome to ''R.sphaeroides'', the ''phaC'' transcription has been found to be reduced under nitrogen deficient condition, although the mechanism of regulation of this gene is not very clear yet..<ref>PMID: 16233588</ref>. in the cited paper, the investigators have included a proposed model for regulation of PHB synthesis in this organism.

5. One group of investigators suggest a role of the genes involved in the nitrogen-related phosphotransferase system (PTS)in PHB accumulation in the organism ''Azotobacter vinelandii''. This group of researchers report that one of the proteins of PTS affect the transcription of the ''phbBAC'' operon(consisting of the ''phbA'', ''phbB'' and ''phbC'' genes) in this organism. Mechanism of this regulation is still unclear.<ref>PMID:17878711</ref>

'''PROBABLE RESEARCH:FOR BETTER UNDERSTANDING OF THIS PROTEIN:CHALLENGE YOUR UNDERSTANDING'''

1. What is the molecular basis of regulation of PHB synthesis in this organism and what is the role of this protein in it?

2. How does environmental factors such as presence or absence of oxygen or nitrogen relate to the activity of this protein in ''R.sphaeroides''?

3. What are the activators or repressor factors, if any, responsible for production of this protein in this organism?

4. Is there a possible role of the nitrogen-related Phosphotransferase system in the regulation of expression of PHB synthase in ''R.sphaeroides''?

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''HOW WAS THE JMOL IMAGE GENERATED?'''

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

'''REFERENCES:'''

<references/>

PAGE CREATED BY MAITREYEE MUKHERJEE, email:maitreyee25@gmail.com

ACKNOWLEDGEMENT: Jill Zeilstra Ryalls, Professor, Bowling Green State University.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-27T14:26:56Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain in Rhodobacter sphaeroides,amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria<ref>PMID: 14999403</ref> which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products<ref>PMID:10416645</ref>. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate <ref> Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of Rhodobacter sphaeroides. Biotechnology Letters. 15:709–714.</ref>.''Rhodobacter sphaeroides'' possesses the Class I phylogenetic group of PHA synthases<ref>PMID:14620841</ref>. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms<ref>PMID:12954080</ref>.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria.

'''PROPERTIES:'''

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/4'>ball and stick model</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/5'>secondary structure</scene>

{{Template:ColorKey_Helix}},
{{Template:ColorKey_Strand}},
{{Template:ColorKey_Turn}}.

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''CURRENT RESEARCH: KNOWLEDGE OF THIS PROTEIN IN ''R.sphaeroides'' AS WELL AS IN OTHER ORGANISMS:'''

1. In ''R.sphaeroides'' FJ1, ''phbC'' has been found to be present on one of two gene locus identified to be involved in the production of PHB. The gene ''phbC'' is present on the same locus along with the other genes responsible for PHB syn thesis such as ''phbZ'', ''phbP'' and ''phbR''. <ref>PMID: 16440119</ref>

2. In a paper by Fales et.al. in 2001, the investigators talk about a possible connection between PHB synthase activity and ''hemA'' transcription in ''R.sphaeroides''. No further investigations answering this have been published after this untill now. The investigators also present a schematic diagram of a proposed dual pathway of PHB synthesis in this organism.<ref>PMID: 11160087</ref>

3. In ''Rhodobacter capsulatus'', ''phaC'' was found to have a constitutive expression in presence or absence of the induction of PHA production, indicating that the control of expression of this gene occurs at the post-translational level in this organism. <ref>PMID: 9251189</ref>

4. In ''Paracoccus denitrificans'', an organism having a close similarity in genome to ''R.sphaeroides'', the ''phaC'' transcription has been found to be reduced under nitrogen deficient condition, although the mechanism of regulation of this gene is not very clear yet..<ref>PMID: 16233588</ref>. in the cited paper, the investigators have included a proposed model for regulation of PHB synthesis in this organism.

5. One group of investigators suggest a role of the genes involved in the nitrogen-related phosphotransferase system (PTS)in PHB accumulation in the organism ''Azotobacter vinelandii''. This group of researchers report that one of the proteins of PTS affect the transcription of the ''phbBAC'' operon(consisting of the ''phbA'', ''phbB'' and ''phbC'' genes) in this organism. Mechanism of this regulation is still unclear.<ref>PMID:17878711</ref>

'''PROBABLE RESEARCH:FOR BETTER UNDERSTANDING OF THIS PROTEIN:CHALLENGE YOUR UNDERSTANDING'''

1. What is the molecular basis of regulation of PHB synthesis in this organism and what is the role of this protein in it?

2. How does environmental factors such as presence or absence of oxygen or nitrogen relate to the activity of this protein in ''R.sphaeroides''?

3. What are the activators or repressor factors, if any, responsible for production of this protein in this organism?

4. Is there a possible role of the nitrogen-related Phosphotransferase system in the regulation of expression of PHB synthase in ''R.sphaeroides''?

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''HOW WAS THE JMOL IMAGE GENERATED?'''

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

'''REFERENCES:'''

<references/>

PAGE CREATED BY MAITREYEE MUKHERJEE, email:maitreyee25@gmail.com

ACKNOWLEDGEMENT: Jill Zeilstra Ryalls, Professor, Bowling Green State University.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-27T00:19:57Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain in Rhodobacter sphaeroides,amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria<ref>PMID: 14999403</ref> which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products<ref>PMID:10416645</ref>. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate <ref> Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of Rhodobacter sphaeroides. Biotechnology Letters. 15:709–714.</ref>.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases<ref>PMID:14620841</ref>. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms<ref>PMID:12954080</ref>.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria.

'''PROPERTIES:'''

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/4'>ball and stick model</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/5'>secondary structure</scene>

{{Template:ColorKey_Helix}},
{{Template:ColorKey_Strand}},
{{Template:ColorKey_Turn}}.

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''CURRENT RESEARCH: KNOWLEDGE OF THIS PROTEIN IN ''R.sphaeroides'' AS WELL AS IN OTHER ORGANISMS:'''

1. In ''R.sphaeroides'' FJ1, ''phbC'' has been found to be present on one of two gene locus identified to be involved in the production of PHB. The gene ''phbC'' is present on the same locus along with the other genes responsible for PHB syn thesis such as ''phbZ'', ''phbP'' and ''phbR''. <ref>PMID: 16440119</ref>

2. In a paper by Fales et.al. in 2001, the investigators talk about a possible connection between PHB synthase activity and ''hemA'' transcription in ''R.sphaeroides''. No further investigations answering this have been published after this untill now. The investigators also present a schematic diagram of a proposed dual pathway of PHB synthesis in this organism.<ref>PMID: 11160087</ref>

3. In ''Rhodobacter capsulatus'', ''phaC'' was found to have a constitutive expression in presence or absence of the induction of PHA production, indicating that the control of expression of this gene occurs at the post-translational level in this organism. <ref>PMID: 9251189</ref>

4. In ''Paracoccus denitrificans'', an organism having a close similarity in genome to ''R.sphaeroides'', the ''phaC'' transcription has been found to be reduced under nitrogen deficient condition, although the mechanism of regulation of this gene is not very clear yet..<ref>PMID: 16233588</ref>. in the cited paper, the investigators have included a proposed model for regulation of PHB synthesis in this organism.

5. One group of investigators suggest a role of the genes involved in the nitrogen-related phosphotransferase system (PTS)in PHB accumulation in the organism ''Azotobacter vinelandii''. This group of researchers report that one of the proteins of PTS affect the transcription of the ''phbBAC'' operon(consisting of the ''phbA'', ''phbB'' and ''phbC'' genes) in this organism. Mechanism of this regulation is still unclear.<ref>PMID:17878711</ref>

'''PROBABLE RESEARCH:FOR BETTER UNDERSTANDING OF THIS PROTEIN:CHALLENGE YOUR UNDERSTANDING'''

1. What is the molecular basis of regulation of PHB synthesis in this organism and what is the role of this protein in it?

2. How does environmental factors such as presence or absence of oxygen or nitrogen relate to the activity of this protein in ''R.sphaeroides''?

3. What are the activators or repressor factors, if any, responsible for production of this protein in this organism?

4. Is there a possible role of the nitrogen-related Phosphotransferase system in the regulation of expression of PHB synthase in ''R.sphaeroides''?

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''HOW WAS THE JMOL IMAGE GENERATED?'''

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

'''REFERENCES:'''

<references/>

PAGE CREATED BY MAITREYEE MUKHERJEE, email:maitreyee25@gmail.com

ACKNOWLEDGEMENT: Jill Zeilstra Ryalls, Professor, Bowling Green State University.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-27T00:19:04Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain in Rhodobacter sphaeroides,amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria<ref>PMID: 14999403</ref> which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products<ref>PMID:10416645</ref>. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate <ref> Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of Rhodobacter sphaeroides. Biotechnology Letters. 15:709–714.</ref>.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases<ref>PMID:14620841</ref>. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms<ref>PMID:12954080</ref>.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria.

'''PROPERTIES:'''

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/4'>ball and stick model</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/5'>secondary structure</scene>

{{Template:ColorKey_Helix}},
{{Template:ColorKey_Strand}},
{{Template:ColorKey_Turn}}.

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''CURRENT RESEARCH: KNOWLEDGE OF THIS PROTEIN IN ''R.sphaeroides'' AS WELL AS IN OTHER ORGANISMS:'''

1. In ''R.sphaeroides'' FJ1, ''phbC'' has been found to be present on one of two gene locus identified to be involved in the production of PHB. The gene ''phbC'' is present on the same locus along with the other genes responsible for PHB syn thesis such as ''phbZ'', ''phbP'' and ''phbR''. <ref>PMID: 16440119</ref>

2. In a paper by Fales et.al. in 2001, the investigators talk about a possible connection between PHB synthase activity and ''hemA'' transcription in ''R.sphaeroides''. No further investigations answering this have been published after this untill now. The investigators also present a schematic diagram of a proposed dual pathway of PHB synthesis in this organism.<ref>PMID: 11160087</ref>

3. In ''Rhodobacter capsulatus'', ''phaC'' was found to have a constitutive expression in presence or absence of the induction of PHA production, indicating that the control of expression of this gene occurs at the post-translational level in this organism. <ref>PMID: 9251189</ref>

4. In ''Paracoccus denitrificans'', an organism having a close similarity in genome to ''R.sphaeroides'', the ''phaC'' transcription has been found to be reduced under nitrogen deficient condition, although the mechanism of regulation of this gene is not very clear yet..<ref>PMID: 16233588</ref>. in the cited paper, the investigators have included a proposed model for regulation of PHB synthesis in this organism.

5. One group of investigators suggest a role of the genes involved in the nitrogen-related phosphotransferase system (PTS)in PHB accumulation in the organism ''Azotobacter vinelandii''. This group of researchers report that one of the proteins of PTS affect the transcription of the ''phbBAC'' operon(consisting of the ''phbA'', ''phbB'' and ''phbC'' genes) in this organism. Mechanism of this regulation is still unclear.<ref>PMID:17878711</ref>

'''PROBABLE RESEARCH:FOR BETTER UNDERSTANDING OF THIS PROTEIN:CHALLENGE YOUR UNDERSTANDING'''

1. What is the molecular basis of regulation of PHB synthesis in this organism and what is the role of this protein in it?

2. How does environmental factors such as presence or absence of oxygen or nitrogen relate to the activity of this protein in ''R.sphaeroides''?

3. What are the activators or repressor factors, if any, responsible for production of this protein in this organism?

4. Is there a possible role of the nitrogen-related Phosphotransferase system in the regulation of expression of PHB synthase in ''R.sphaeroides''?

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''HOW WAS THE JMOL IMAGE GENERATED?'''

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

'''REFERENCES:'''

<references/>

PAGE CREATED BY MAITREYEE MUKHERJEE, email:maitreyee25@gmail

ACKNOWLEDGEMENT: Jill Zeilstra Ryalls, Professor, Bowling Green State University.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T23:58:53Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain in Rhodobacter sphaeroides,amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria<ref>PMID: 14999403</ref> which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products<ref>PMID:10416645</ref>. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate <ref> Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of Rhodobacter sphaeroides. Biotechnology Letters. 15:709–714.</ref>.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases<ref>PMID:14620841</ref>. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms<ref>PMID:12954080</ref>.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria.

'''PROPERTIES:'''

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/4'>ball and stick model</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/5'>secondary structure</scene>

{{Template:ColorKey_Helix}},
{{Template:ColorKey_Strand}},
{{Template:ColorKey_Turn}}.

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''CURRENT RESEARCH: KNOWLEDGE OF THIS PROTEIN IN ''R.sphaeroides'' AS WELL AS OTHER ORGANISMS:'''

1. In ''R.sphaeroides'' FJ1, ''phbC'' has been found to be present on one of two gene locus identified to be involved in the production of PHB. The gene ''phbC'' is present on the same locus along with the other genes responsible for PHB syn thesis such as ''phbZ'', ''phbP'' and ''phbR''. <ref>PMID: 16440119</ref>

2. In a paper by Fales et.al. in 2001, the investigators talk about a possible connection between PHB synthase activity and ''hemA'' transcription in ''R.sphaeroides''. No further investigations answering this have been published after this untill now. The investigators also present a schematic diagram of a proposed dual pathway of PHB synthesis in this organism.<ref>PMID: 11160087</ref>

3. In ''Rhodobacter capsulatus'', ''phaC'' was found to have a constitutive expression in presence or absence of the induction of PHA production, indicating that the control of expression of this gene occurs at the post-translational level in this organism. <ref>PMID: 9251189</ref>

4. In ''Paracoccus denitrificans'', an organism having a close similarity in genome to ''R.sphaeroides'', the ''phaC'' transcription has been found to be reduced under nitrogen deficient condition, although the mechanism of regulation of this gene is not very clear yet..<ref>PMID: 16233588</ref>. in the cited paper, the investigators have included a proposed model for regulation of PHB synthesis in this organism.

5. One group of investigators suggest a role of the genes involved in the nitrogen-related phosphotransferase system (PTS)in PHB accumulation in the organism ''Azotobacter vinelandii''. This group of researchers report that one of the proteins of PTS affect the transcription of the ''phbBAC'' operon(consisting of the ''phbA'', ''phbB'' and ''phbC'' genes) in this organism. Mechanism of this regulation is still unclear.<ref>PMID:17878711</ref>

'''PROBABLE RESEARCH:FOR BETTER UNDERSTANDING OF THIS PROTEIN:CHALLENGE YOUR UNDERSTANDING'''

1. What is the molecular basis of regulation of PHB synthesis in this organism and what is the role of this protein in it?

2. How does environmental factors such as presence or absence of oxygen or nitrogen relate to the activity of this protein in ''R.sphaeroides''?

3. What are the activators or repressor factors, if any, responsible for production of this protein in this organism?

4. Is there a possible role of the nitrogen-related Phosphotransferase system in the regulation of expression of PHB synthase in ''R.sphaeroides''?

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''HOW WAS THE JMOL IMAGE GENERATED?'''

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

'''REFERENCES:'''

<references/>

PAGE CREATED BY MAITREYEE MUKHERJEE, email:maitreyee25@gmail

ACKNOWLEDGEMENT: Jill Zeilstra Ryalls, Professor, Bowling Green State University.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T23:40:06Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain in Rhodobacter sphaeroides,amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria<ref>PMID: 14999403</ref> which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products<ref>PMID:10416645</ref>. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate <ref> Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of Rhodobacter sphaeroides. Biotechnology Letters. 15:709–714.</ref>.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases<ref>PMID:14620841</ref>. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms<ref>PMID:12954080</ref>.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/4'>ball and stick model</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/5'>secondary structure</scene>

{{Template:ColorKey_Helix}},
{{Template:ColorKey_Strand}},
{{Template:ColorKey_Turn}}.

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''CURRENT RESEARCH: KNOWLEDGE OF THIS PROTEIN IN ''R.sphaeroides'' AS WELL AS OTHER ORGANISMS:'''

1. In ''R.sphaeroides'' FJ1, ''phbC'' has been found to be present on one of two gene locus identified to be involved in the production of PHB. The gene ''phbC'' is present on the same locus along with the other genes responsible for PHB syn thesis such as ''phbZ'', ''phbP'' and ''phbR''. <ref>PMID: 16440119</ref>

2. In a paper by Fales et.al. in 2001, the investigators talk about a possible connection between PHB synthase activity and ''hemA'' transcription in ''R.sphaeroides''. No further investigations answering this have been published after this untill now. The investigators also present a schematic diagram of a proposed dual pathway of PHB synthesis in this organism.<ref>PMID: 11160087</ref>

3. In ''Rhodobacter capsulatus'', ''phaC'' was found to have a constitutive expression in presence or absence of the induction of PHA production, indicating that the control of expression of this gene occurs at the post-translational level in this organism. <ref>PMID: 9251189</ref>

4. In ''Paracoccus denitrificans'', an organism having a close similarity in genome to ''R.sphaeroides'', the ''phaC'' transcription has been found to be reduced under nitrogen deficient condition, although the mechanism of regulation of this gene is not very clear yet..<ref>PMID: 16233588</ref>. in the cited paper, the investigators have included a proposed model for regulation of PHB synthesis in this organism.

'''PROBABLE RESEARCH:FOR BETTER UNDERSTANDING OF THIS PROTEIN:CHALLENGE YOUR UNDERSTANDING'''

1. What is the molecular basis of regulation of PHB synthesis in this organism and what is the role of this protein in it?

2. How does environmental factors such as presence or absence of oxygen or nitrogen relate to the activity of this protein in ''R.sphaeroides''?

3. What are the activators or repressor factors, if any, responsible for production of this protein in this organism?

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''HOW WAS THE JMOL IMAGE GENERATED?'''

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

'''REFERENCES:'''

<references/>

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' expression in ''Rhodobacter sphaeroides'' 2.4.1: effect of a transposon insertion in the ''hbdA'' gene. Journal of Bacteriology. 183:1568–1576.

2. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, molecular analysis, and expression of the polyhydroxyalkanoic acid synthase (''phaC'') gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate production in ''Rhodobacter'' ''capsulatus'': genes, mutants, expression, and physiology. Applied and Environmental Microbiology. 63:3003–3009.

5. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-hydroxybutyrate (PHB) biosynthesis in ''Alcaligenes eutrophus'' H16, identification and characterization of the PHB polymerase gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

6. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25:3-19.

7. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochemical Journal. 376:15–33.

8. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) homeostasis: the role of PHA synthase. Natural Product Reports. 20:445-457.

9. Tae-Kwon, K. J. Young-Mi. M. Tri Vo., S.Suteaki., and L.Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress.22: 1541-1546.

10. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular analysis of the poly(3-hydroxyalkanoate) synthase gene from a methylotrophic bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T23:38:19Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain in Rhodobacter sphaeroides,amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria<ref>PMID: 14999403</ref> which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products<ref>PMID:10416645</ref>. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate <ref> Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of Rhodobacter sphaeroides. Biotechnology Letters. 15:709–714.</ref>.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases<ref>PMID:14620841</ref>. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms<ref>PMID:12954080</ref>.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/4'>ball and stick model</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/5'>secondary structure</scene>

{{Template:ColorKey_Helix}},
{{Template:ColorKey_Strand}},
{{Template:ColorKey_Turn}}.

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''CURRENT RESEARCH: KNOWLEDGE OF THIS PROTEIN IN ''R.sphaeroides'' AS WELL AS OTHER ORGANISMS:'''

1. In ''R.sphaeroides'' FJ1, ''phbC'' has been found to be present on one of two gene locus identified to be involved in the production of PHB. The gene ''phbC'' is present on the same locus along with the other genes responsible for PHB syn thesis such as ''phbZ'', ''phbP'' and ''phbR''. <ref>PMID: 16440119</ref>

2. In a paper by Fales et.al. in 2001, the investigators talk about a possible connection between PHB synthase activity and ''hemA'' transcription in ''R.sphaeroides''. No further investigations answering this have been published after this untill now. The investigators also present a schematic diagram of a proposed dual pathway of PHB synthesis in this organism.<ref>PMID: 11160087</ref>

3. In ''Rhodobacter capsulatus'', ''phaC'' was found to have a constitutive expression in presence or absence of the induction of PHA production, indicating that the control of expression of this gene occurs at the post-translational level in this organism. <ref>PMID: 9251189</ref>

4. In ''Paracoccus denitrificans'', an organism having a close similarity in genome to ''R.sphaeroides'', the ''phaC'' transcription has been found to be reduced under nitrogen deficient condition, although the mechanism of regulation of this gene is not very clear yet..<ref>PMID: 16233588</ref>

'''PROBABLE RESEARCH:FOR BETTER UNDERSTANDING OF THIS PROTEIN:CHALLENGE YOUR UNDERSTANDING'''

1. What is the molecular basis of regulation of PHB synthesis in this organism and what is the role of this protein in it?

2. How does environmental factors such as presence or absence of oxygen or nitrogen relate to the activity of this protein in ''R.sphaeroides''?

3. What are the activators or repressor factors, if any, responsible for production of this protein in this organism?

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''HOW WAS THE JMOL IMAGE GENERATED?'''

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

'''REFERENCES:'''

<references/>

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' expression in ''Rhodobacter sphaeroides'' 2.4.1: effect of a transposon insertion in the ''hbdA'' gene. Journal of Bacteriology. 183:1568–1576.

2. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, molecular analysis, and expression of the polyhydroxyalkanoic acid synthase (''phaC'') gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate production in ''Rhodobacter'' ''capsulatus'': genes, mutants, expression, and physiology. Applied and Environmental Microbiology. 63:3003–3009.

5. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-hydroxybutyrate (PHB) biosynthesis in ''Alcaligenes eutrophus'' H16, identification and characterization of the PHB polymerase gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

6. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25:3-19.

7. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochemical Journal. 376:15–33.

8. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) homeostasis: the role of PHA synthase. Natural Product Reports. 20:445-457.

9. Tae-Kwon, K. J. Young-Mi. M. Tri Vo., S.Suteaki., and L.Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress.22: 1541-1546.

10. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular analysis of the poly(3-hydroxyalkanoate) synthase gene from a methylotrophic bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T23:16:05Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain in Rhodobacter sphaeroides,amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria<ref>PMID: 14999403</ref> which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products<ref>PMID:10416645</ref>. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate <ref> Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of Rhodobacter sphaeroides. Biotechnology Letters. 15:709–714.</ref>.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases<ref>PMID:14620841</ref>. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms<ref>PMID:12954080</ref>.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/4'>ball and stick model</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/5'>secondary structure</scene>

{{Template:ColorKey_Helix}},
{{Template:ColorKey_Strand}},
{{Template:ColorKey_Turn}}.

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''CURRENT RESEARCH: KNOWLEDGE OF THIS PROTEIN IN ''R.sphaeroides'' AS WELL AS OTHER ORGANISMS:'''

1. In ''R.sphaeroides'' FJ1, ''phbC'' has been found to be present on one of two gene locus identified to be involved in the production of PHB. The gene ''phbC'' is present on the same locus along with the other genes responsible for PHB syn thesis such as ''phbZ'', ''phbP'' and ''phbR''. <ref>PMID: 16440119</ref>

2. In a paper by Fales et.al. in 2001, the investigators talk about a possible connection between PHB synthase activity and ''hemA'' transcription in ''R.sphaeroides''. The investigators also present a schematic diagram of a proposed dual pathway of PHB synthesis in this organism.<ref>PMID: 11160087</ref> No further investigations have been published regarding this untill now.

'''PROBABLE RESEARCH:FOR BETTER UNDERSTANDING OF THIS PROTEIN:CHALLENGE YOUR UNDERSTANDING'''

1. What is the molecular basis of regulation of PHB synthesis in this organism and what is the role of this protein in it?

2. How does environmental factors such as presence or absence of oxygen or nitrogen relate to the activity of this protein in ''R.sphaeroides''?

3. What are the activators or repressor factors, if any, responsible for production of this protein in this organism?

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''HOW WAS THE JMOL IMAGE GENERATED?'''

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

'''REFERENCES:'''

<references/>

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' expression in ''Rhodobacter sphaeroides'' 2.4.1: effect of a transposon insertion in the ''hbdA'' gene. Journal of Bacteriology. 183:1568–1576.

2. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, molecular analysis, and expression of the polyhydroxyalkanoic acid synthase (''phaC'') gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate production in ''Rhodobacter'' ''capsulatus'': genes, mutants, expression, and physiology. Applied and Environmental Microbiology. 63:3003–3009.

5. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-hydroxybutyrate (PHB) biosynthesis in ''Alcaligenes eutrophus'' H16, identification and characterization of the PHB polymerase gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

6. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25:3-19.

7. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochemical Journal. 376:15–33.

8. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) homeostasis: the role of PHA synthase. Natural Product Reports. 20:445-457.

9. Tae-Kwon, K. J. Young-Mi. M. Tri Vo., S.Suteaki., and L.Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress.22: 1541-1546.

10. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular analysis of the poly(3-hydroxyalkanoate) synthase gene from a methylotrophic bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T23:03:07Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain in Rhodobacter sphaeroides,amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria<ref>PMID: 14999403</ref> which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products<ref>PMID:10416645</ref>. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate<ref>PMID: 11160087</ref> <ref> Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of Rhodobacter sphaeroides. Biotechnology Letters. 15:709–714.</ref>.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases<ref>PMID:14620841</ref>. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms<ref>PMID:12954080</ref>.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/4'>ball and stick model</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/5'>secondary structure</scene>

{{Template:ColorKey_Helix}},
{{Template:ColorKey_Strand}},
{{Template:ColorKey_Turn}}.

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''CURRENT RESEARCH: KNOWLEDGE OF THIS PROTEIN IN ''R.sphaeroides'' AS WELL AS OTHER ORGANISMS:'''

1. In ''R.sphaeroides'' FJ1, ''phbC'' has been found to be present on one of two gene locus identified to be involved in the production of PHB. The gene ''phbC'' is present on the same locus along with the other genes responsible for PHB syn thesis such as ''phbZ'', ''phbP'' and ''phbR''. <ref>PMID: 16440119</ref>
'''PROBABLE RESEARCH:FOR BETTER UNDERSTANDING OF THIS PROTEIN:CHALLENGE YOUR UNDERSTANDING'''

1. What is the molecular basis of regulation of PHB synthesis in this organism and what is the role of this protein in it?

2. How does environmental factors such as presence or absence of oxygen or nitrogen relate to the activity of this protein in ''R.sphaeroides''?

3. What are the activators or repressor factors, if any, responsible for production of this protein in this organism?

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''HOW WAS THE JMOL IMAGE GENERATED?'''

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

'''REFERENCES:'''

<references/>

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' expression in ''Rhodobacter sphaeroides'' 2.4.1: effect of a transposon insertion in the ''hbdA'' gene. Journal of Bacteriology. 183:1568–1576.

2. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, molecular analysis, and expression of the polyhydroxyalkanoic acid synthase (''phaC'') gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate production in ''Rhodobacter'' ''capsulatus'': genes, mutants, expression, and physiology. Applied and Environmental Microbiology. 63:3003–3009.

5. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-hydroxybutyrate (PHB) biosynthesis in ''Alcaligenes eutrophus'' H16, identification and characterization of the PHB polymerase gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

6. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25:3-19.

7. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochemical Journal. 376:15–33.

8. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) homeostasis: the role of PHA synthase. Natural Product Reports. 20:445-457.

9. Tae-Kwon, K. J. Young-Mi. M. Tri Vo., S.Suteaki., and L.Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress.22: 1541-1546.

10. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular analysis of the poly(3-hydroxyalkanoate) synthase gene from a methylotrophic bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T22:53:17Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain in Rhodobacter sphaeroides,amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria<ref>PMID: 14999403</ref> which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products<ref>PMID:10416645</ref>. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate<ref>PMID: 11160087</ref> <ref> Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of Rhodobacter sphaeroides. Biotechnology Letters. 15:709–714.</ref>.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases<ref>PMID:14620841</ref>. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms<ref>PMID:12954080</ref>.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/4'>ball and stick model</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/5'>secondary structure</scene>

{{Template:ColorKey_Helix}},
{{Template:ColorKey_Strand}},
{{Template:ColorKey_Turn}}.

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''CURRENT RESEARCH: KNOWLEDGE OF THIS PROTEIN IN ''R.sphaeroides'' AS WELL AS OTHER ORGANISMS:'''

'''PROBABLE RESEARCH:FOR BETTER UNDERSTANDING OF THIS PROTEIN:CHALLENGE YOUR UNDERSTANDING'''

1. What is the molecular basis of regulation of PHB synthesis in this organism and what is the role of this protein in it?

2. How does environmental factors such as presence or absence of oxygen or nitrogen relate to the activity of this protein in ''R.sphaeroides''?

3. What are the activators or repressor factors, if any, responsible for production of this protein in this organism?

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''HOW WAS THE JMOL IMAGE GENERATED?'''

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

'''REFERENCES:'''

<references/>

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' expression in ''Rhodobacter sphaeroides'' 2.4.1: effect of a transposon insertion in the ''hbdA'' gene. Journal of Bacteriology. 183:1568–1576.

2. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, molecular analysis, and expression of the polyhydroxyalkanoic acid synthase (''phaC'') gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate production in ''Rhodobacter'' ''capsulatus'': genes, mutants, expression, and physiology. Applied and Environmental Microbiology. 63:3003–3009.

5. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-hydroxybutyrate (PHB) biosynthesis in ''Alcaligenes eutrophus'' H16, identification and characterization of the PHB polymerase gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

6. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25:3-19.

7. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochemical Journal. 376:15–33.

8. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) homeostasis: the role of PHA synthase. Natural Product Reports. 20:445-457.

9. Tae-Kwon, K. J. Young-Mi. M. Tri Vo., S.Suteaki., and L.Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress.22: 1541-1546.

10. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular analysis of the poly(3-hydroxyalkanoate) synthase gene from a methylotrophic bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T22:49:50Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain in Rhodobacter sphaeroides,amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria<ref>PMID: 14999403</ref> which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products<ref>PMID:10416645</ref>. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate<ref>PMID: 11160087</ref> <ref> Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of Rhodobacter sphaeroides. Biotechnology Letters. 15:709–714.</ref>.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases<ref>PMID:14620841</ref>. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms<ref>PMID:12954080</ref>.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/4'>ball and stick model</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/5'>secondary structure</scene>

{{Template:ColorKey_Helix}},
{{Template:ColorKey_Strand}},
{{Template:ColorKey_Turn}}.

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''PROBABLE RESEARCH:FOR BETTER UNDERSTANDING OF THIS PROTEIN:CHALLENGE YOUR UNDERSTANDING'''

1. What is the molecular basis of regulation of PHB synthesis in this organism and what is the role of this protein in it?

2. How does environmental factors such as presence or absence of oxygen or nitrogen relate to the activity of this protein in ''R.sphaeroides''?

3. What are the activators or repressor factors, if any, responsible for production of this protein in this organism?

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''HOW WAS THE JMOL IMAGE GENERATED?'''

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

'''REFERENCES:'''

<references/>

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' expression in ''Rhodobacter sphaeroides'' 2.4.1: effect of a transposon insertion in the ''hbdA'' gene. Journal of Bacteriology. 183:1568–1576.

2. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, molecular analysis, and expression of the polyhydroxyalkanoic acid synthase (''phaC'') gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate production in ''Rhodobacter'' ''capsulatus'': genes, mutants, expression, and physiology. Applied and Environmental Microbiology. 63:3003–3009.

5. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-hydroxybutyrate (PHB) biosynthesis in ''Alcaligenes eutrophus'' H16, identification and characterization of the PHB polymerase gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

6. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25:3-19.

7. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochemical Journal. 376:15–33.

8. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) homeostasis: the role of PHA synthase. Natural Product Reports. 20:445-457.

9. Tae-Kwon, K. J. Young-Mi. M. Tri Vo., S.Suteaki., and L.Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress.22: 1541-1546.

10. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular analysis of the poly(3-hydroxyalkanoate) synthase gene from a methylotrophic bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T22:48:20Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain in Rhodobacter sphaeroides,amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria<ref>PMID: 14999403</ref> which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products<ref>PMID:10416645</ref>. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate<ref>PMID: 11160087</ref> <ref> Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of Rhodobacter sphaeroides. Biotechnology Letters. 15:709–714.</ref>.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases<ref>PMID:14620841</ref>. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms<ref>PMID:12954080</ref>.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/4'>ball and stick model</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/5'>secondary structure</scene>

{{Template:ColorKey_Helix}},
{{Template:ColorKey_Strand}},
{{Template:ColorKey_Turn}}.

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''PROBABLE RESEARCH:FOR BETTER UNDERSTANDING OF THIS PROTEIN:'''

1. What is the molecular basis of regulation of PHB synthesis in this organism and what is the role of this protein in it?

2. How does environmental factors such as presence or absence of oxygen or nitrogen relate to the activity of this protein in ''R.sphaeroides''?

3. What are the activators or repressor factors, if any, responsible for production of this protein in this organism?

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''HOW WAS THE JMOL IMAGE GENERATED?'''

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

'''REFERENCES:'''

<references/>

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' expression in ''Rhodobacter sphaeroides'' 2.4.1: effect of a transposon insertion in the ''hbdA'' gene. Journal of Bacteriology. 183:1568–1576.

2. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, molecular analysis, and expression of the polyhydroxyalkanoic acid synthase (''phaC'') gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate production in ''Rhodobacter'' ''capsulatus'': genes, mutants, expression, and physiology. Applied and Environmental Microbiology. 63:3003–3009.

5. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-hydroxybutyrate (PHB) biosynthesis in ''Alcaligenes eutrophus'' H16, identification and characterization of the PHB polymerase gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

6. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25:3-19.

7. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochemical Journal. 376:15–33.

8. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) homeostasis: the role of PHA synthase. Natural Product Reports. 20:445-457.

9. Tae-Kwon, K. J. Young-Mi. M. Tri Vo., S.Suteaki., and L.Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress.22: 1541-1546.

10. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular analysis of the poly(3-hydroxyalkanoate) synthase gene from a methylotrophic bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T21:57:23Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain in Rhodobacter sphaeroides,amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria<ref>PMID: 14999403</ref> which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products<ref>PMID:10416645</ref>. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate<ref>PMID: 11160087</ref> <ref> Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of Rhodobacter sphaeroides. Biotechnology Letters. 15:709–714.</ref>.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases<ref>PMID:14620841</ref>. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms<ref>PMID:12954080</ref>.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/4'>ball and stick model</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/5'>secondary structure</scene>

{{Template:ColorKey_Helix}},
{{Template:ColorKey_Strand}},
{{Template:ColorKey_Turn}}.

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''HOW WAS THE JMOL IMAGE GENERATED?'''

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

'''REFERENCES:'''

<references/>

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' expression in ''Rhodobacter sphaeroides'' 2.4.1: effect of a transposon insertion in the ''hbdA'' gene. Journal of Bacteriology. 183:1568–1576.

2. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, molecular analysis, and expression of the polyhydroxyalkanoic acid synthase (''phaC'') gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate production in ''Rhodobacter'' ''capsulatus'': genes, mutants, expression, and physiology. Applied and Environmental Microbiology. 63:3003–3009.

5. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-hydroxybutyrate (PHB) biosynthesis in ''Alcaligenes eutrophus'' H16, identification and characterization of the PHB polymerase gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

6. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25:3-19.

7. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochemical Journal. 376:15–33.

8. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) homeostasis: the role of PHA synthase. Natural Product Reports. 20:445-457.

9. Tae-Kwon, K. J. Young-Mi. M. Tri Vo., S.Suteaki., and L.Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress.22: 1541-1546.

10. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular analysis of the poly(3-hydroxyalkanoate) synthase gene from a methylotrophic bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T21:29:35Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain in Rhodobacter sphaeroides,amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria<ref>PMID: 14999403</ref> which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products<ref>PMID:10416645</ref>. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate<ref>PMID: 11160087</ref> <ref> Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of Rhodobacter sphaeroides. Biotechnology Letters. 15:709–714.</ref>.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases<ref>PMID:14620841</ref>. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms<ref>PMID:12954080</ref>.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA<ref>PMID: 12954080</ref>. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/4'>ball and stick model</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/5'>secondary structure</scene>

{{Template:ColorKey_Helix}},
{{Template:ColorKey_Strand}},
{{Template:ColorKey_Turn}}.

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''HOW WAS THE JMOL IMAGE GENERATED?'''

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

'''REFERENCES:'''

<references/>

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' expression in ''Rhodobacter sphaeroides'' 2.4.1: effect of a transposon insertion in the ''hbdA'' gene. Journal of Bacteriology. 183:1568–1576.

2. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, molecular analysis, and expression of the polyhydroxyalkanoic acid synthase (''phaC'') gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate production in ''Rhodobacter'' ''capsulatus'': genes, mutants, expression, and physiology. Applied and Environmental Microbiology. 63:3003–3009.

5. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-hydroxybutyrate (PHB) biosynthesis in ''Alcaligenes eutrophus'' H16, identification and characterization of the PHB polymerase gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

6. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25:3-19.

7. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochemical Journal. 376:15–33.

8. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) homeostasis: the role of PHA synthase. Natural Product Reports. 20:445-457.

9. Tae-Kwon, K. J. Young-Mi. M. Tri Vo., S.Suteaki., and L.Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress.22: 1541-1546.

10. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular analysis of the poly(3-hydroxyalkanoate) synthase gene from a methylotrophic bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T21:23:39Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain in Rhodobacter sphaeroides,amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products<ref>PMID:10416645</ref>. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate<ref>PMID: 11160087</ref> <ref> Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of Rhodobacter sphaeroides. Biotechnology Letters. 15:709–714.</ref>.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms<ref>PMID:12954080</ref>.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA<ref>PMID: 12954080</ref>. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/4'>ball and stick model</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/5'>secondary structure</scene>

{{Template:ColorKey_Helix}},
{{Template:ColorKey_Strand}},
{{Template:ColorKey_Turn}}.

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''HOW WAS THE JMOL IMAGE GENERATED?'''

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

'''REFERENCES:'''

<references/>

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' expression in ''Rhodobacter sphaeroides'' 2.4.1: effect of a transposon insertion in the ''hbdA'' gene. Journal of Bacteriology. 183:1568–1576.

2. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, molecular analysis, and expression of the polyhydroxyalkanoic acid synthase (''phaC'') gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate production in ''Rhodobacter'' ''capsulatus'': genes, mutants, expression, and physiology. Applied and Environmental Microbiology. 63:3003–3009.

5. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-hydroxybutyrate (PHB) biosynthesis in ''Alcaligenes eutrophus'' H16, identification and characterization of the PHB polymerase gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

6. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25:3-19.

7. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochemical Journal. 376:15–33.

8. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) homeostasis: the role of PHA synthase. Natural Product Reports. 20:445-457.

9. Tae-Kwon, K. J. Young-Mi. M. Tri Vo., S.Suteaki., and L.Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress.22: 1541-1546.

10. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular analysis of the poly(3-hydroxyalkanoate) synthase gene from a methylotrophic bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T21:20:34Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain in Rhodobacter sphaeroides,amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate<ref>PMID: 11160087</ref> <ref> Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of Rhodobacter sphaeroides. Biotechnology Letters. 15:709–714.</ref>.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms<ref>PMID:12954080</ref>.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA<ref>PMID: 12954080</ref>. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/4'>ball and stick model</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/5'>secondary structure</scene>

{{Template:ColorKey_Helix}},
{{Template:ColorKey_Strand}},
{{Template:ColorKey_Turn}}.

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''HOW WAS THE JMOL IMAGE GENERATED?'''

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

'''REFERENCES:'''

<references/>

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' expression in ''Rhodobacter sphaeroides'' 2.4.1: effect of a transposon insertion in the ''hbdA'' gene. Journal of Bacteriology. 183:1568–1576.

2. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, molecular analysis, and expression of the polyhydroxyalkanoic acid synthase (''phaC'') gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate production in ''Rhodobacter'' ''capsulatus'': genes, mutants, expression, and physiology. Applied and Environmental Microbiology. 63:3003–3009.

5. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-hydroxybutyrate (PHB) biosynthesis in ''Alcaligenes eutrophus'' H16, identification and characterization of the PHB polymerase gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

6. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25:3-19.

7. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochemical Journal. 376:15–33.

8. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) homeostasis: the role of PHA synthase. Natural Product Reports. 20:445-457.

9. Tae-Kwon, K. J. Young-Mi. M. Tri Vo., S.Suteaki., and L.Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress.22: 1541-1546.

10. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular analysis of the poly(3-hydroxyalkanoate) synthase gene from a methylotrophic bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T19:38:00Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain in Rhodobacter sphaeroides,amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/4'>ball and stick model</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/5'>secondary structure</scene>

{{Template:ColorKey_Helix}},
{{Template:ColorKey_Strand}},
{{Template:ColorKey_Turn}}.

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''HOW WAS THE JMOL IMAGE GENERATED?'''

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

'''REFERENCES:'''

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' expression in ''Rhodobacter sphaeroides'' 2.4.1: effect of a transposon insertion in the ''hbdA'' gene. Journal of Bacteriology. 183:1568–1576.

2. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, molecular analysis, and expression of the polyhydroxyalkanoic acid synthase (''phaC'') gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate production in ''Rhodobacter'' ''capsulatus'': genes, mutants, expression, and physiology. Applied and Environmental Microbiology. 63:3003–3009.

5. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-hydroxybutyrate (PHB) biosynthesis in ''Alcaligenes eutrophus'' H16, identification and characterization of the PHB polymerase gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

6. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25:3-19.

7. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochemical Journal. 376:15–33.

8. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) homeostasis: the role of PHA synthase. Natural Product Reports. 20:445-457.

9. Tae-Kwon, K. J. Young-Mi. M. Tri Vo., S.Suteaki., and L.Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress.22: 1541-1546.

10. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular analysis of the poly(3-hydroxyalkanoate) synthase gene from a methylotrophic bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T19:36:37Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain in ''Rhodobacter sphaeroides'', amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/4'>ball and stick model</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/5'>secondary structure</scene>

{{Template:ColorKey_Helix}},
{{Template:ColorKey_Strand}},
{{Template:ColorKey_Turn}}.

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''HOW WAS THE JMOL IMAGE GENERATED?'''

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

'''REFERENCES:'''

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' expression in ''Rhodobacter sphaeroides'' 2.4.1: effect of a transposon insertion in the ''hbdA'' gene. Journal of Bacteriology. 183:1568–1576.

2. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, molecular analysis, and expression of the polyhydroxyalkanoic acid synthase (''phaC'') gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate production in ''Rhodobacter'' ''capsulatus'': genes, mutants, expression, and physiology. Applied and Environmental Microbiology. 63:3003–3009.

5. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-hydroxybutyrate (PHB) biosynthesis in ''Alcaligenes eutrophus'' H16, identification and characterization of the PHB polymerase gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

6. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25:3-19.

7. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochemical Journal. 376:15–33.

8. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) homeostasis: the role of PHA synthase. Natural Product Reports. 20:445-457.

9. Tae-Kwon, K. J. Young-Mi. M. Tri Vo., S.Suteaki., and L.Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress.22: 1541-1546.

10. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular analysis of the poly(3-hydroxyalkanoate) synthase gene from a methylotrophic bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T19:33:41Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain, amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/4'>ball and stick model</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/5'>secondary structure</scene>

{{Template:ColorKey_Helix}},
{{Template:ColorKey_Strand}},
{{Template:ColorKey_Turn}}.

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''HOW WAS THE JMOL IMAGE GENERATED?'''

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

'''REFERENCES:'''

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' expression in ''Rhodobacter sphaeroides'' 2.4.1: effect of a transposon insertion in the ''hbdA'' gene. Journal of Bacteriology. 183:1568–1576.

2. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, molecular analysis, and expression of the polyhydroxyalkanoic acid synthase (''phaC'') gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate production in ''Rhodobacter'' ''capsulatus'': genes, mutants, expression, and physiology. Applied and Environmental Microbiology. 63:3003–3009.

5. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-hydroxybutyrate (PHB) biosynthesis in ''Alcaligenes eutrophus'' H16, identification and characterization of the PHB polymerase gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

6. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25:3-19.

7. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochemical Journal. 376:15–33.

8. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) homeostasis: the role of PHA synthase. Natural Product Reports. 20:445-457.

9. Tae-Kwon, K. J. Young-Mi. M. Tri Vo., S.Suteaki., and L.Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress.22: 1541-1546.

10. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular analysis of the poly(3-hydroxyalkanoate) synthase gene from a methylotrophic bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T19:25:17Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain, amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/4'>ball and stick model</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''HOW WAS THE JMOL IMAGE GENERATED?'''

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

'''REFERENCES:'''

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' expression in ''Rhodobacter sphaeroides'' 2.4.1: effect of a transposon insertion in the ''hbdA'' gene. Journal of Bacteriology. 183:1568–1576.

2. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, molecular analysis, and expression of the polyhydroxyalkanoic acid synthase (''phaC'') gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate production in ''Rhodobacter'' ''capsulatus'': genes, mutants, expression, and physiology. Applied and Environmental Microbiology. 63:3003–3009.

5. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-hydroxybutyrate (PHB) biosynthesis in ''Alcaligenes eutrophus'' H16, identification and characterization of the PHB polymerase gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

6. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25:3-19.

7. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochemical Journal. 376:15–33.

8. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) homeostasis: the role of PHA synthase. Natural Product Reports. 20:445-457.

9. Tae-Kwon, K. J. Young-Mi. M. Tri Vo., S.Suteaki., and L.Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress.22: 1541-1546.

10. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular analysis of the poly(3-hydroxyalkanoate) synthase gene from a methylotrophic bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T19:21:38Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain, amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase domain in N>C rainbow color</scene>

{{Template:ColorKey_N2CRainbow}}

{{Template:ColorKey_Amino2CarboxyRainbow}}

{{Template:ColorKey_523Rainbow}}

{{Template:ColorKey_N52C3Rainbow}}

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase domain showing hydrophobic/polar residues</scene>

{{Template:ColorKey_Hydrophobic}}, {{Template:ColorKey_Polar}}

'''Number of amino acids:''' 601

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''HOW WAS THE JMOL IMAGE GENERATED?'''

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

'''REFERENCES:'''

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' expression in ''Rhodobacter sphaeroides'' 2.4.1: effect of a transposon insertion in the ''hbdA'' gene. Journal of Bacteriology. 183:1568–1576.

2. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, molecular analysis, and expression of the polyhydroxyalkanoic acid synthase (''phaC'') gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate production in ''Rhodobacter'' ''capsulatus'': genes, mutants, expression, and physiology. Applied and Environmental Microbiology. 63:3003–3009.

5. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-hydroxybutyrate (PHB) biosynthesis in ''Alcaligenes eutrophus'' H16, identification and characterization of the PHB polymerase gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

6. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25:3-19.

7. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochemical Journal. 376:15–33.

8. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) homeostasis: the role of PHA synthase. Natural Product Reports. 20:445-457.

9. Tae-Kwon, K. J. Young-Mi. M. Tri Vo., S.Suteaki., and L.Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress.22: 1541-1546.

10. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular analysis of the poly(3-hydroxyalkanoate) synthase gene from a methylotrophic bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T19:13:41Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain, amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

'''Number of amino acids:''' 601

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase showing hydrophobic/polar residues</scene>

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''HOW WAS THE JMOL IMAGE GENERATED?'''

1. First retrieve your protein sequence from http://www.ncbi.nlm.nih.gov/.

2. Go to 3D-JIGSAW page http://bmm.cancerresearchuk.org/~3djigsaw/ and paste the sequence on the submission page. A .pdb format image of your protein will be sent to you on your email which can be opened by RASMOL.

3. Upload this file on Proteopedia and then load the JMol applet for the protein following instructions on the Help:Editing page http://www.proteopedia.org/wiki/index.php/Help:Editing.

4. You can edit your protein by using the scene authoring tools after loading the applet.

'''REFERENCES:'''

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' expression in ''Rhodobacter sphaeroides'' 2.4.1: effect of a transposon insertion in the ''hbdA'' gene. Journal of Bacteriology. 183:1568–1576.

2. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, molecular analysis, and expression of the polyhydroxyalkanoic acid synthase (''phaC'') gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate production in ''Rhodobacter'' ''capsulatus'': genes, mutants, expression, and physiology. Applied and Environmental Microbiology. 63:3003–3009.

5. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-hydroxybutyrate (PHB) biosynthesis in ''Alcaligenes eutrophus'' H16, identification and characterization of the PHB polymerase gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

6. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25:3-19.

7. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochemical Journal. 376:15–33.

8. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) homeostasis: the role of PHA synthase. Natural Product Reports. 20:445-457.

9. Tae-Kwon, K. J. Young-Mi. M. Tri Vo., S.Suteaki., and L.Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress.22: 1541-1546.

10. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular analysis of the poly(3-hydroxyalkanoate) synthase gene from a methylotrophic bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T18:23:42Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain, amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

'''Number of amino acids:''' 601

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase showing hydrophobic/polar residues</scene>

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''HOW WAS THE JMOL IMAGE GENERATED?'''

'''REFERENCES:'''

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' expression in ''Rhodobacter sphaeroides'' 2.4.1: effect of a transposon insertion in the ''hbdA'' gene. Journal of Bacteriology. 183:1568–1576.

2. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, molecular analysis, and expression of the polyhydroxyalkanoic acid synthase (''phaC'') gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate production in ''Rhodobacter'' ''capsulatus'': genes, mutants, expression, and physiology. Applied and Environmental Microbiology. 63:3003–3009.

5. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-hydroxybutyrate (PHB) biosynthesis in ''Alcaligenes eutrophus'' H16, identification and characterization of the PHB polymerase gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

6. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25:3-19.

7. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochemical Journal. 376:15–33.

8. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) homeostasis: the role of PHA synthase. Natural Product Reports. 20:445-457.

9. Tae-Kwon, K. J. Young-Mi. M. Tri Vo., S.Suteaki., and L.Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress.22: 1541-1546.

10. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular analysis of the poly(3-hydroxyalkanoate) synthase gene from a methylotrophic bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T17:38:35Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain, amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

'''Number of amino acids:''' 601

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase showing hydrophobic/polar residues</scene>

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be generated in pixels(here the size is 600), then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''HOW WAS THE JMOL IMAGE GENERATED?'''

'''REFERENCES:'''

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' expression in ''Rhodobacter sphaeroides'' 2.4.1: effect of a transposon insertion in the ''hbdA'' gene. Journal of Bacteriology. 183:1568–1576."PMID:"11160087

2. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, molecular analysis, and expression of the polyhydroxyalkanoic acid synthase (''phaC'') gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate production in ''Rhodobacter'' ''capsulatus'': genes, mutants, expression, and physiology. Applied and Environmental Microbiology. 63:3003–3009.

5. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-hydroxybutyrate (PHB) biosynthesis in ''Alcaligenes eutrophus'' H16, identification and characterization of the PHB polymerase gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

6. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25:3-19.

7. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochemical Journal. 376:15–33.

8. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) homeostasis: the role of PHA synthase. Natural Product Reports. 20:445-457.

9. Tae-Kwon, K. J. Young-Mi. M. Tri Vo., S.Suteaki., and L.Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress.22: 1541-1546.

10. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular analysis of the poly(3-hydroxyalkanoate) synthase gene from a methylotrophic bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T17:35:30Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain, amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

'''Number of amino acids:''' 601

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase showing hydrophobic/polar residues</scene>

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be genrated in pixels, then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''HOW WAS THE JMOL IMAGE GENERATED?'''

'''REFERENCES:'''

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' expression in ''Rhodobacter sphaeroides'' 2.4.1: effect of a transposon insertion in the ''hbdA'' gene. Journal of Bacteriology. 183:1568–1576."PMID:"11160087

2. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, molecular analysis, and expression of the polyhydroxyalkanoic acid synthase (''phaC'') gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate production in ''Rhodobacter'' ''capsulatus'': genes, mutants, expression, and physiology. Applied and Environmental Microbiology. 63:3003–3009.

5. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-hydroxybutyrate (PHB) biosynthesis in ''Alcaligenes eutrophus'' H16, identification and characterization of the PHB polymerase gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

6. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25:3-19.

7. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochemical Journal. 376:15–33.

8. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) homeostasis: the role of PHA synthase. Natural Product Reports. 20:445-457.

9. Tae-Kwon, K. J. Young-Mi. M. Tri Vo., S.Suteaki., and L.Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress.22: 1541-1546.

10. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular analysis of the poly(3-hydroxyalkanoate) synthase gene from a methylotrophic bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T17:33:26Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain, amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

'''Number of amino acids:''' 601

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase showing hydrophobic/polar residues</scene>

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html,

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be genrated in pixels, then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''HOW WAS THE JMOL IMAGE GENERATED?'''

'''REFERENCES:'''

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' expression in ''Rhodobacter sphaeroides'' 2.4.1: effect of a transposon insertion in the ''hbdA'' gene. Journal of Bacteriology. 183:1568–1576."PMID:"11160087

2. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, molecular analysis, and expression of the polyhydroxyalkanoic acid synthase (''phaC'') gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate production in ''Rhodobacter'' ''capsulatus'': genes, mutants, expression, and physiology. Applied and Environmental Microbiology. 63:3003–3009.

5. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-hydroxybutyrate (PHB) biosynthesis in ''Alcaligenes eutrophus'' H16, identification and characterization of the PHB polymerase gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

6. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25:3-19.

7. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochemical Journal. 376:15–33.

8. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) homeostasis: the role of PHA synthase. Natural Product Reports. 20:445-457.

9. Tae-Kwon, K. J. Young-Mi. M. Tri Vo., S.Suteaki., and L.Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress.22: 1541-1546.

10. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular analysis of the poly(3-hydroxyalkanoate) synthase gene from a methylotrophic bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T17:19:33Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain, amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

'''Number of amino acids:''' 601

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase showing hydrophobic/polar residues</scene>

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''HOW WAS THE JMOL IMAGE GENERATED?'''

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

1. Go to the POLYVIEW 3D homepage, http://polyview.cchmc.org/polyview3d.html,

2. On the submission form, first select 'animation' in the "type of request" section, select the size of the animation to be genrated in pixels, then upload the PDB format protein structure file in the "source of structural data" section.

3. On the "chain color and rendering section" select 'cartoon' and 'secondary structure'.

4. On "advanced structural annotation" section select 'docking models in Capri format'.

5. Any other forms for the animation may be selected by referring to the "Samples" according to the protein structure to be animated.

'''REFERENCES:'''

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' expression in ''Rhodobacter sphaeroides'' 2.4.1: effect of a transposon insertion in the ''hbdA'' gene. Journal of Bacteriology. 183:1568–1576.

2. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, molecular analysis, and expression of the polyhydroxyalkanoic acid synthase (''phaC'') gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate production in ''Rhodobacter'' ''capsulatus'': genes, mutants, expression, and physiology. Applied and Environmental Microbiology. 63:3003–3009.

5. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-hydroxybutyrate (PHB) biosynthesis in ''Alcaligenes eutrophus'' H16, identification and characterization of the PHB polymerase gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

6. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25:3-19.

7. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochemical Journal. 376:15–33.

8. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) homeostasis: the role of PHA synthase. Natural Product Reports. 20:445-457.

9. Tae-Kwon, K. J. Young-Mi. M. Tri Vo., S.Suteaki., and L.Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress.22: 1541-1546.

10. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular analysis of the poly(3-hydroxyalkanoate) synthase gene from a methylotrophic bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T17:06:05Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain, amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

'''Number of amino acids:''' 601

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase showing hydrophobic/polar residues</scene>

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''HOW WAS THE JMOL IMAGE GENERATED?'''

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

'''REFERENCES:'''

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' expression in ''Rhodobacter sphaeroides'' 2.4.1: effect of a transposon insertion in the ''hbdA'' gene. Journal of Bacteriology. 183:1568–1576.

2. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, molecular analysis, and expression of the polyhydroxyalkanoic acid synthase (''phaC'') gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate production in ''Rhodobacter'' ''capsulatus'': genes, mutants, expression, and physiology. Applied and Environmental Microbiology. 63:3003–3009.

5. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-hydroxybutyrate (PHB) biosynthesis in ''Alcaligenes eutrophus'' H16, identification and characterization of the PHB polymerase gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

6. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25:3-19.

7. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochemical Journal. 376:15–33.

8. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) homeostasis: the role of PHA synthase. Natural Product Reports. 20:445-457.

9. Tae-Kwon, K. J. Young-Mi. M. Tri Vo., S.Suteaki., and L.Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress.22: 1541-1546.

10. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular analysis of the poly(3-hydroxyalkanoate) synthase gene from a methylotrophic bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T17:04:55Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain, amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

'''Number of amino acids:''' 601

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase showing hydrophobic/polar residues</scene>

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''HOW WAS THE JMOL IMAGE GENERATED?'''

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

'''REFERENCES:'''

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' expression in ''Rhodobacter sphaeroides'' 2.4.1: effect of a transposon insertion in the ''hbdA'' gene. Journal of Bacteriology. 183:1568–1576.

2. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, molecular analysis, and expression of the polyhydroxyalkanoic acid synthase (''phaC'') gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate production in ''Rhodobacter'' ''capsulatus'': genes, mutants, expression, and physiology. Applied and Environmental Microbiology. 63:3003–3009.

5. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-hydroxybutyrate (PHB) biosynthesis in ''Alcaligenes eutrophus'' H16, identification and characterization of the PHB polymerase gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

6. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25:3-19.

7. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochemical Journal. 376:15–33.

8. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) homeostasis: the role of PHA synthase. Natural Product Reports. 20:445-457.

9. Tae-Kwon, K. J. Young-Mi. M. Tri Vo., S.Suteaki., and L.Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress.22: 1541-1546.

10. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular analysis of the poly(3-hydroxyalkanoate) synthase gene from a methylotrophic bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T17:03:44Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain, amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

'''Number of amino acids:''' 601

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase showing hydrophobic/polar residues</scene>

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''HOW WAS THE JMOL IMAGE GENERATED?'''

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

'''REFERENCES:'''

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of hemA expression in ''Rhodobacter sphaeroides'' 2.4.1: effect of a transposon insertion in the hbdA gene. Journal of Bacteriology. 183:1568–1576.

2. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, molecular analysis, and expression of the polyhydroxyalkanoic acid synthase (''phaC'') gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate production in ''Rhodobacter'' ''capsulatus'': genes, mutants, expression, and physiology. Applied and Environmental Microbiology. 63:3003–3009.

5. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-hydroxybutyrate (PHB) biosynthesis in ''Alcaligenes eutrophus'' H16, identification and characterization of the PHB polymerase gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

6. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25:3-19.

7. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochemical Journal. 376:15–33.

8. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) homeostasis: the role of PHA synthase. Natural Product Reports. 20:445-457.

9. Tae-Kwon, K. J. Young-Mi. M. Tri Vo., S.Suteaki., and L.Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress.22: 1541-1546.

10. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular analysis of the poly(3-hydroxyalkanoate) synthase gene from a methylotrophic bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T16:44:06Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain, amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products. PHB synthase encoded by ''phaC'' is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

'''Number of amino acids:''' 601

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase showing hydrophobic/polar residues</scene>

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''HOW WAS THE JMOL IMAGE GENERATED?'''

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

'''REFERENCES:'''

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' Expression in ''Rhodobacter sphaeroides'' 2.4.1: Effect of a Transposon Insertion in the ''hbdA'' Gene. Journal of Bacteriology. 183:1568–1576.

2. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate Production in ''Rhodobacter'' ''capsulatus'': Genes, Mutants, Expression, and Physiology. Applied and Environmental Microbiology. 63:3003–3009.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, Molecular Analysis, and Expression of the Polyhydroxyalkanoic Acid Synthase (''phaC'') Gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular Analysis of the Poly(3-Hydroxyalkanoate) Synthase Gene from a Methylotrophic Bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

5. Kadouri, D., S. Burdman., E. Jurkevitch., and Y. Okon. 2002. Identification and Isolation of Genes Involved in Poly ß-Hydroxybutyrate Biosynthesis in ''Azospirillum brasilense'' and Characterization of a ''phbC'' Mutant. Applied and Environmental Microbiology. 68(6): 2943–2949.

6. Tae-Kwon, K. J. Young-Mi.. M. Tri Vo., S. Suteaki., and L. Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress. 22:1541-1546.

7. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25: 3-19.

8. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-Hydroxybutyrate (PHB) Biosynthesis In ''Alcaligenes eutrophus'' H16, Identification And Characterization of the PHB Polymerase Gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

9. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) Homeostasis: The Role of the PHA Synthase. Nat. Prod. Rep. 20:445 – 457.

10. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochem. J. 376:15–33.

11. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T16:34:44Z

Maitreyee Mukherjee:

<applet load='FILE_consurf1240241051_pipe.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain, amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products. PHB synthase or phaC is the gene that is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

'''Number of amino acids:''' 601

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase showing hydrophobic/polar residues</scene>

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''HOW WAS THE JMOL IMAGE GENERATED?'''

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

'''REFERENCES:'''

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' Expression in ''Rhodobacter sphaeroides'' 2.4.1: Effect of a Transposon Insertion in the ''hbdA'' Gene. Journal of Bacteriology. 183:1568–1576.

2. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate Production in ''Rhodobacter'' ''capsulatus'': Genes, Mutants, Expression, and Physiology. Applied and Environmental Microbiology. 63:3003–3009.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, Molecular Analysis, and Expression of the Polyhydroxyalkanoic Acid Synthase (''phaC'') Gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular Analysis of the Poly(3-Hydroxyalkanoate) Synthase Gene from a Methylotrophic Bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

5. Kadouri, D., S. Burdman., E. Jurkevitch., and Y. Okon. 2002. Identification and Isolation of Genes Involved in Poly ß-Hydroxybutyrate Biosynthesis in ''Azospirillum brasilense'' and Characterization of a ''phbC'' Mutant. Applied and Environmental Microbiology. 68(6): 2943–2949.

6. Tae-Kwon, K. J. Young-Mi.. M. Tri Vo., S. Suteaki., and L. Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress. 22:1541-1546.

7. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25: 3-19.

8. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-Hydroxybutyrate (PHB) Biosynthesis In ''Alcaligenes eutrophus'' H16, Identification And Characterization of the PHB Polymerase Gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

9. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) Homeostasis: The Role of the PHA Synthase. Nat. Prod. Rep. 20:445 – 457.

10. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochem. J. 376:15–33.

11. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T16:12:18Z

Maitreyee Mukherjee:

<applet load='Phb_synthase_1.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain, amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products. PHB synthase or phaC is the gene that is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

'''Number of amino acids:''' 601

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase showing hydrophobic/polar residues</scene>

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE DOMAIN SHOWING AMINO ACIDS 286 TO 547
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''HOW WAS THE JMOL IMAGE GENERATED?'''

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

'''REFERENCES:'''

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' Expression in ''Rhodobacter sphaeroides'' 2.4.1: Effect of a Transposon Insertion in the ''hbdA'' Gene. Journal of Bacteriology. 183:1568–1576.

2. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate Production in ''Rhodobacter'' ''capsulatus'': Genes, Mutants, Expression, and Physiology. Applied and Environmental Microbiology. 63:3003–3009.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, Molecular Analysis, and Expression of the Polyhydroxyalkanoic Acid Synthase (''phaC'') Gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular Analysis of the Poly(3-Hydroxyalkanoate) Synthase Gene from a Methylotrophic Bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

5. Kadouri, D., S. Burdman., E. Jurkevitch., and Y. Okon. 2002. Identification and Isolation of Genes Involved in Poly ß-Hydroxybutyrate Biosynthesis in ''Azospirillum brasilense'' and Characterization of a ''phbC'' Mutant. Applied and Environmental Microbiology. 68(6): 2943–2949.

6. Tae-Kwon, K. J. Young-Mi.. M. Tri Vo., S. Suteaki., and L. Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress. 22:1541-1546.

7. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25: 3-19.

8. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-Hydroxybutyrate (PHB) Biosynthesis In ''Alcaligenes eutrophus'' H16, Identification And Characterization of the PHB Polymerase Gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

9. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) Homeostasis: The Role of the PHA Synthase. Nat. Prod. Rep. 20:445 – 457.

10. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochem. J. 376:15–33.

11. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T15:50:15Z

Maitreyee Mukherjee:

<applet load='Phb_synthase_1.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain, amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products. PHB synthase or phaC is the gene that is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

'''Number of amino acids:''' 601

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase showing hydrophobic/polar residues</scene>

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''HOW WAS THE JMOL IMAGE GENERATED?'''

'''HOW WAS THE ANIMATED IMAGE GENERATED?'''

'''REFERENCES:'''

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' Expression in ''Rhodobacter sphaeroides'' 2.4.1: Effect of a Transposon Insertion in the ''hbdA'' Gene. Journal of Bacteriology. 183:1568–1576.

2. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate Production in ''Rhodobacter'' ''capsulatus'': Genes, Mutants, Expression, and Physiology. Applied and Environmental Microbiology. 63:3003–3009.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, Molecular Analysis, and Expression of the Polyhydroxyalkanoic Acid Synthase (''phaC'') Gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular Analysis of the Poly(3-Hydroxyalkanoate) Synthase Gene from a Methylotrophic Bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

5. Kadouri, D., S. Burdman., E. Jurkevitch., and Y. Okon. 2002. Identification and Isolation of Genes Involved in Poly ß-Hydroxybutyrate Biosynthesis in ''Azospirillum brasilense'' and Characterization of a ''phbC'' Mutant. Applied and Environmental Microbiology. 68(6): 2943–2949.

6. Tae-Kwon, K. J. Young-Mi.. M. Tri Vo., S. Suteaki., and L. Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress. 22:1541-1546.

7. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25: 3-19.

8. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-Hydroxybutyrate (PHB) Biosynthesis In ''Alcaligenes eutrophus'' H16, Identification And Characterization of the PHB Polymerase Gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

9. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) Homeostasis: The Role of the PHA Synthase. Nat. Prod. Rep. 20:445 – 457.

10. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochem. J. 376:15–33.

11. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T15:47:54Z

Maitreyee Mukherjee:

<applet load='Phb_synthase_1.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain, amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products. PHB synthase or phaC is the gene that is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

'''Number of amino acids:''' 601

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase showing hydrophobic/polar residues</scene>

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data1.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''HOW WAS THE JMOL IMAGE GENERATED?'''

'''REFERENCES:'''

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' Expression in ''Rhodobacter sphaeroides'' 2.4.1: Effect of a Transposon Insertion in the ''hbdA'' Gene. Journal of Bacteriology. 183:1568–1576.

2. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate Production in ''Rhodobacter'' ''capsulatus'': Genes, Mutants, Expression, and Physiology. Applied and Environmental Microbiology. 63:3003–3009.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, Molecular Analysis, and Expression of the Polyhydroxyalkanoic Acid Synthase (''phaC'') Gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular Analysis of the Poly(3-Hydroxyalkanoate) Synthase Gene from a Methylotrophic Bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

5. Kadouri, D., S. Burdman., E. Jurkevitch., and Y. Okon. 2002. Identification and Isolation of Genes Involved in Poly ß-Hydroxybutyrate Biosynthesis in ''Azospirillum brasilense'' and Characterization of a ''phbC'' Mutant. Applied and Environmental Microbiology. 68(6): 2943–2949.

6. Tae-Kwon, K. J. Young-Mi.. M. Tri Vo., S. Suteaki., and L. Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress. 22:1541-1546.

7. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25: 3-19.

8. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-Hydroxybutyrate (PHB) Biosynthesis In ''Alcaligenes eutrophus'' H16, Identification And Characterization of the PHB Polymerase Gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

9. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) Homeostasis: The Role of the PHA Synthase. Nat. Prod. Rep. 20:445 – 457.

10. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochem. J. 376:15–33.

11. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

File:Get data1.gif

2009-04-26T15:45:04Z

Maitreyee Mukherjee:

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T15:34:37Z

Maitreyee Mukherjee:

<applet load='Phb_synthase_1.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain, amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products. PHB synthase or phaC is the gene that is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

'''Number of amino acids:''' 601

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase showing hydrophobic/polar residues</scene>

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''HOW WAS THE JMOL IMAGE GENERATED?'''

'''REFERENCES:'''

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' Expression in ''Rhodobacter sphaeroides'' 2.4.1: Effect of a Transposon Insertion in the ''hbdA'' Gene. Journal of Bacteriology. 183:1568–1576.

2. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate Production in ''Rhodobacter'' ''capsulatus'': Genes, Mutants, Expression, and Physiology. Applied and Environmental Microbiology. 63:3003–3009.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, Molecular Analysis, and Expression of the Polyhydroxyalkanoic Acid Synthase (''phaC'') Gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular Analysis of the Poly(3-Hydroxyalkanoate) Synthase Gene from a Methylotrophic Bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

5. Kadouri, D., S. Burdman., E. Jurkevitch., and Y. Okon. 2002. Identification and Isolation of Genes Involved in Poly ß-Hydroxybutyrate Biosynthesis in ''Azospirillum brasilense'' and Characterization of a ''phbC'' Mutant. Applied and Environmental Microbiology. 68(6): 2943–2949.

6. Tae-Kwon, K. J. Young-Mi.. M. Tri Vo., S. Suteaki., and L. Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress. 22:1541-1546.

7. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25: 3-19.

8. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-Hydroxybutyrate (PHB) Biosynthesis In ''Alcaligenes eutrophus'' H16, Identification And Characterization of the PHB Polymerase Gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

9. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) Homeostasis: The Role of the PHA Synthase. Nat. Prod. Rep. 20:445 – 457.

10. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochem. J. 376:15–33.

11. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.

User:Maitreyee Mukherjee/Sandbox 1

2009-04-26T15:27:47Z

Maitreyee Mukherjee:

<applet load='Phb_synthase_1.pdb' size='500' frame='true' align='right' caption='Putative PHB synthase domain, amino acids 286 to 547' />

'''INTRODUCTION:'''

''Rhodobacter sphaeroides'' is a metabolically versatile purple photosynthetic α-proteobacteria which produces polyhydroxybutyrate (PHB) as inclusions inside its cell. PHB is a biopolymer which has the properties similar to synthetic polymers such as polyethylene and hence has a potential of being used as a biopolymer. The biochemical pathway for PHB production in this organism is complex and is controlled by the actions of several gene products. PHB synthase or phaC is the gene that is responsible for conversion of the monomeric precursor of PHB R(-)-β-Hydroxybutyryl-CoA into polyhydroxybutyrate.Rhodobacter sphaeroides possesses the Class I phylogenetic group of PHA synthases. Class I PHA synthases prefer utilization of CoA thioesters of various (R)-3-hydroxy fatty acids comprising of 3 to 5 carbon atoms.

'''FUNCTION:'''

The enzyme catalyses the conversion of the monomeric precursor of PHB into PHB with the release of CoA. In other words these enzymes mediate the conversion of a soluble substrate into polymerized insoluble inclusions inside the cells of these bacteria. Upon covalent catalysis of polyester chain formation, this soluble enzyme gets converted to amphipathic enzyme (The amphipathic enzyme conjugates are analogous to certain membrane-bound proteins such as cytochrome b, which have been shown to comprise a hydrophilic protein core anchored to a lipid bilayer by a hydrophobic polypeptide tail). Initiation of a self assembly process occurs and this results in the formation of the insoluble cytoplasmic inclusions with a phospholipids monolayer and covalently attached polyester synthases at the surface.

'''PROPERTIES:'''

'''Number of amino acids:''' 601

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/1'>PHB sythase in N>C rainbow color</scene>

<scene name='User:Maitreyee_Mukherjee/Sandbox_1/Putative_phb_synthase/3'>PHB synthase showing hydrophobic/polar residues</scene>

'''Molecular weight:''' 66831.6

'''Theoretical pI:''' 5.63

'''Amino acid sequence:'''

1 mateeqspgs grdaqferln anltridels krltaaltkr klsdpalhgp sgdvflkamt

61 aymaemmqnp akilehqisf wgkslkhyve aqhqlvkgel kpppdvtpkd rrfsnplwqt

121 hpffnylkqq ylmnaeavnq avealehiep sdkkrveyfs rqivdlfspt nffgtnpdal

181 eraiatdges lvqglenlvr dieanngdll vtladpeafq vgqnlatteg svvyrnrmfe

241 liqykpttet vhetpllifp pwinkfyild lkpqnsllkw lvdqgftvfv vswvnpdksy

301 agigmddyir egymramaev rsitrqkqin avgyciagtt ltltlahlqk agdpsvrsat

361 ffttltdfsd pgevgvflnd dfvdgierqv avdgildktf msrtfsylrs ndliyqpaik

421 symmgeappa fdllywngdg tnlpaqmave ylrglcqqdr laggtfpvlg spvglkdvtl

481 pvcaiacetd hiapwkssfn gfrqfgstdk tfilsqsghv agivnppsrn kyghytnegp

541 agtpesfreg aefhagswwp rwgawlaers gkqvparqpg dskhpelapa pgsyvaavgg

601 a

'''Amino acid Composition:'''

Ala (A) 52 8.7%,
Arg (R) 29 4.8%,
Asn (N) 26 4.3%,
Asp (D) 34 5.7%,
Cys (C) 4 0.7%,
Gln (Q) 29 4.8%,
Glu (E) 35 5.8%,
Gly (G) 46 7.7%,
His (H) 13 2.2%,
Ile (I) 24 4.0%,
Leu (L) 54 9.0%,
Lys (K) 28 4.7%,
Met (M) 14 2.3%,
Phe (F) 31 5.2%,
Pro (P) 40 6.7%,
Ser (S) 36 6.0%,
Thr (T) 37 6.2%,
Trp (W) 11 1.8%,
Tyr (Y) 20 3.3%.
Val (V) 38 6.3%
Pyl (O) 0 0.0%
Sec (U) 0 0.0%

Total number of negatively charged residues (Asp + Glu): 69
Total number of positively charged residues (Arg + Lys): 57

'''Atomic composition:'''

Carbon C 3004
Hydrogen H 4612
Nitrogen N 808
Oxygen O 888
Sulfur S 18

'''Total number of atoms:''' 9330

'''Extinction coefficients:'''

Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.

Ext. coefficient 90550
Abs 0.1% (=1 g/l) 1.355, assuming ALL Cys residues appear as half cystines
Ext. coefficient 90300
Abs 0.1% (=1 g/l) 1.351, assuming NO Cys residues appear as half cystines

'''Estimated half-life:'''

The N-terminal of the sequence considered is M (Met).

The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).
>20 hours (yeast, in vivo).
>10 hours (''Escherichia coli'', in vivo).

'''Instability index:'''

The instability index (II) is 41.97
This classifies the protein as unstable.

'''Aliphatic index:''' 77.60

'''Grand average of hydropathicity (GRAVY):''' -0.330

[[Image:Get_data.gif|alt text]]

'''ANIMATED IMAGE OF THE PUTATIVE PHB SYNTHASE
'''IN''' '''''Rhodobacter sphaeroides''''' '''OBTAINED FROM POLYVIEW-3D'''''

'''REFERENCES:'''

1. Fales, L., L. Kryszak., and J. Zeilstra-Ryalls. 2001. Control of ''hemA'' Expression in ''Rhodobacter sphaeroides'' 2.4.1: Effect of a Transposon Insertion in the ''hbdA'' Gene. Journal of Bacteriology. 183:1568–1576.

2. Kranz, R.G., K. K. Gabbert., T. A. Locke., and M. T. Madigan. 1997. Polyhydroxyalkanoate Production in ''Rhodobacter'' ''capsulatus'': Genes, Mutants, Expression, and Physiology. Applied and Environmental Microbiology. 63:3003–3009.

3. Kolibachuk, D., A. Miller., and D. Dennis. 1999. Cloning, Molecular Analysis, and Expression of the Polyhydroxyalkanoic Acid Synthase (''phaC'') Gene from ''Chromobacterium violaceum''. Applied and Environmental Microbiology. 65:3561–3565.

4. Ueda, S., T. Yabutani., A. Maehara., and T. Yamane. 1996. Molecular Analysis of the Poly(3-Hydroxyalkanoate) Synthase Gene from a Methylotrophic Bacterium, ''Paracoccus denitrificans''. Journal of Bacteriology. 178:774–779.

5. Kadouri, D., S. Burdman., E. Jurkevitch., and Y. Okon. 2002. Identification and Isolation of Genes Involved in Poly ß-Hydroxybutyrate Biosynthesis in ''Azospirillum brasilense'' and Characterization of a ''phbC'' Mutant. Applied and Environmental Microbiology. 68(6): 2943–2949.

6. Tae-Kwon, K. J. Young-Mi.. M. Tri Vo., S. Suteaki., and L. Yong-Hyun. 2006. Metabolic engineering and characterization of ''phaC1'' and ''phaC2'' genes from ''Pseudomonas putida'' KCTC1639 for overproduction of medium-chain-length polyhydroxyalkanoate. Biotechnology Progress. 22:1541-1546.

7. Rehm, H. A. B., and A. Steinbüchel. 1999. Biochemical and genetic analysis of PHA synthases and other proteins required for PHA synthesis. International Journal of Biological Macromolecules. 25: 3-19.

8. Peoples, P.O., and A. J. Sinskey. 1989. Poly-ß-Hydroxybutyrate (PHB) Biosynthesis In ''Alcaligenes eutrophus'' H16, Identification And Characterization of the PHB Polymerase Gene (''phbC''). The Journal of Biological Chemistry. 264(26):15298-15303.

9. Stubbe, J. A., and J. Tian. 2003. Polyhydroxyalkanoate (PHA) Homeostasis: The Role of the PHA Synthase. Nat. Prod. Rep. 20:445 – 457.

10. Rehm, Bernd H. A. B. 2003. Polyester synthases: natural catalysts for plastics. Biochem. J. 376:15–33.

11. Hustede, E., and A. Steinbuchel. 1993. Characterization of the polyhydroxyalkanoate synthase gene locus of ''Rhodobacter'' ''sphaeroides''. Biotechnology Letters. 15:709–714.