Sandbox Reserved 717: Difference between revisions

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==The 33kDa Fragment <ref>PMID: 12888344</ref> <ref>PMID:11530935</ref>==
==='''The 33kDa Fragment <ref>PMID: 12888344</ref> <ref>PMID:11530935</ref>'''===
The 33kDa fragment (PDB:1H6W)  was generated in the presence of EDTA. This fragment contains the residues <scene name='Sandbox_Reserved_717/Myfirstscene/1'>85-395</scene> and <scene name='Sandbox_Reserved_717/Mysecondscene/1'>518-527</scene> .  The residues 397-517 are lacking because of internal deletion.
The 33kDa fragment (PDB:1H6W)  was generated in the presence of EDTA. This fragment contains the residues <scene name='Sandbox_Reserved_717/Myfirstscene/1'>85-395</scene> and <scene name='Sandbox_Reserved_717/Mysecondscene/1'>518-527</scene> .  The residues 397-517 are lacking because of internal deletion.
The 33kDa fragment can be further sub-divided into two subunits. The <scene name='Sandbox_Reserved_717/Mythirdscene/1'>neck</scene>  (residue 333-341) and the collar (residues <scene name='Sandbox_Reserved_717/Mythirdscene/2'>342-396 </scene> plus <scene name='Sandbox_Reserved_717/Mythirdscene/3'> 518-527</scene>). The neck connects the body of the fibre to its C-terminal collar and receptor binding-site. It consists of a triple alpha-helix which is built by the residues 333-341.
The 33kDa fragment can be further sub-divided into two subunits. The <scene name='Sandbox_Reserved_717/Mythirdscene/1'>neck</scene>  (residue 333-341) and the collar (residues <scene name='Sandbox_Reserved_717/Mythirdscene/2'>342-396 </scene> plus <scene name='Sandbox_Reserved_717/Mythirdscene/3'> 518-527</scene>). The neck connects the body of the fibre to its C-terminal collar and receptor binding-site. It consists of a triple alpha-helix which is built by the residues 333-341.
The collar domain is a small globular domain. It contains six beta-strands and an alpha-helix.
The collar domain is a small globular domain. It contains six beta-strands and an alpha-helix.


==The 45kDa Fragment <ref> doi:10.1038/nsb970</ref> ==
==='''The 45kDa Fragment <ref> doi:10.1038/nsb970</ref>''' ===
For generating the 45kDa fragment the full length gp12 was co-expressed with its chaperone gp57 and purified [1]. Like the 33kDa fragment it also starts with the amino acid Leu85. The 45kDa fragment contains the residues <scene name='Sandbox_Reserved_717/Mythirdscene/4'>397-517</scene> , which are in the 33kDa fragment internal deleted.
For generating the 45kDa fragment the full length gp12 was co-expressed with its chaperone gp57 and purified [1]. Like the 33kDa fragment it also starts with the amino acid Leu85. The 45kDa fragment contains the residues <scene name='Sandbox_Reserved_717/Mythirdscene/4'>397-517</scene> , which are in the 33kDa fragment internal deleted.
Like the 33kDa fragment the 45kDa fragment can also be divided into two subunits. These two subunits are called head (residues <scene name='Sandbox_Reserved_717/Mythirdscene/5'>397-446 </scene> and residues <scene name='Sandbox_Reserved_717/Mythirdscene/6'>487-517</scene> ) and <scene name='Sandbox_Reserved_717/Mythirdscene/7'>bonnet</scene>  (residues 447-487). On the border between the head and the bonnet subunit there is a metal-binding site.   
Like the 33kDa fragment the 45kDa fragment can also be divided into two subunits. These two subunits are called head (residues <scene name='Sandbox_Reserved_717/Mythirdscene/5'>397-446 </scene> and residues <scene name='Sandbox_Reserved_717/Mythirdscene/6'>487-517</scene> ) and <scene name='Sandbox_Reserved_717/Mythirdscene/7'>bonnet</scene>  (residues 447-487). On the border between the head and the bonnet subunit there is a metal-binding site.   


==Receptor-Binding Domain <ref> doi:10.1038/nsb970</ref> ==
==='''Receptor-Binding Domain <ref> doi:10.1038/nsb970</ref>''' ===
Gp12 is fixed with its N-terminal domain to the baseplate. So the C-terminal domain has to be involved in LPS-binding. To detect where the receptor-binding domain is, full-length gp12, 33kDa fragments and 45kDa fragments were immobilised in micro-plate wells and were allowed to bind to bacteria[1]. The result was that the 33kDa fragment did never bind to a bacteria. The 45kDa fragment did bind. So the receptor-binding domain is absent in the 33kDa fragment but present in the 45kDa fragment. The residues which are present in the 45kDa fragment and lacking in the 33kDa fragment are the residues <scene name='Sandbox_Reserved_717/Mythirdscene/8'>397-517</scene>. They are referred to be part of the receptor-binding domain.
Gp12 is fixed with its N-terminal domain to the baseplate. So the C-terminal domain has to be involved in LPS-binding. To detect where the receptor-binding domain is, full-length gp12, 33kDa fragments and 45kDa fragments were immobilised in micro-plate wells and were allowed to bind to bacteria[1]. The result was that the 33kDa fragment did never bind to a bacteria. The 45kDa fragment did bind. So the receptor-binding domain is absent in the 33kDa fragment but present in the 45kDa fragment. The residues which are present in the 45kDa fragment and lacking in the 33kDa fragment are the residues <scene name='Sandbox_Reserved_717/Mythirdscene/8'>397-517</scene>. They are referred to be part of the receptor-binding domain.
The receptor-binding domain can be sub-divided into head and bonnet. On the border between these two subdomains there is a metal-binding site. This site binds presumably to zinc. <scene name='Sandbox_Reserved_717/Mynewscene/3'>Two His amino acids </scene> (<scene name='Sandbox_Reserved_717/Mynewscene/1'>His445</scene> and <scene name='Sandbox_Reserved_717/Mynewscene/2'>His447</scene>) from each monomere are octahedrally coordinated around the zinc.  
The receptor-binding domain can be sub-divided into head and bonnet. On the border between these two subdomains there is a metal-binding site. This site binds presumably to zinc. <scene name='Sandbox_Reserved_717/Mynewscene/3'>Two His amino acids </scene> (<scene name='Sandbox_Reserved_717/Mynewscene/1'>His445</scene> and <scene name='Sandbox_Reserved_717/Mynewscene/2'>His447</scene>) from each monomere are octahedrally coordinated around the zinc.  
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The receptor-binding domain can be compared to a flower bud. This flower bud has got 12 petals which are organised in a 3-fold symmetry. At the bottom there are the residues 406-432 they form the  <scene name='Sandbox_Reserved_717/Mynewscene/4'>first petal </scene> , just above there are the residues 489-504 which form the <scene name='Sandbox_Reserved_717/Mynewscene/5'>second petal</scene> . The <scene name='Sandbox_Reserved_717/Mynewscene/6'>third petal </scene> is formed by the residues 450-470 and at the top there are the residues 470-480 and form the <scene name='Sandbox_Reserved_717/Mynewscene/7'>fourth petal</scene> . The complete and active receptor-binding domain is built by the trimeric protein. This trimeric proteine structure is stabilised by many inter-domain hydrogen bounds. These hydrogen bounds can have the profiles: main-chain-main-chain, main-chain-side-chain and side-chain-side-chain.
The receptor-binding domain can be compared to a flower bud. This flower bud has got 12 petals which are organised in a 3-fold symmetry. At the bottom there are the residues 406-432 they form the  <scene name='Sandbox_Reserved_717/Mynewscene/4'>first petal </scene> , just above there are the residues 489-504 which form the <scene name='Sandbox_Reserved_717/Mynewscene/5'>second petal</scene> . The <scene name='Sandbox_Reserved_717/Mynewscene/6'>third petal </scene> is formed by the residues 450-470 and at the top there are the residues 470-480 and form the <scene name='Sandbox_Reserved_717/Mynewscene/7'>fourth petal</scene> . The complete and active receptor-binding domain is built by the trimeric protein. This trimeric proteine structure is stabilised by many inter-domain hydrogen bounds. These hydrogen bounds can have the profiles: main-chain-main-chain, main-chain-side-chain and side-chain-side-chain.


==The LPS-Bindind Site <ref> doi:10.1038/nsb970</ref>==
==='''The LPS-Bindind Site <ref> doi:10.1038/nsb970</ref>'''===
The exact position of the lipo-polysaccharide(LPS)-binding site is not known. Proteolysis experiments showed, that it contains to the domain with the residues <scene name='Sandbox_Reserved_717/Mythirdscene/8'>397-517</scene>. Compared with the homologous bacteriophage T4-like strain AR1 sequence, which also binds to the same LPS core molecule like gp12, there can be some possible binding residues be assumed.  
The exact position of the lipo-polysaccharide(LPS)-binding site is not known. Proteolysis experiments showed, that it contains to the domain with the residues <scene name='Sandbox_Reserved_717/Mythirdscene/8'>397-517</scene>. Compared with the homologous bacteriophage T4-like strain AR1 sequence, which also binds to the same LPS core molecule like gp12, there can be some possible binding residues be assumed.  


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==Ligands and their Binding-Sites==
=='''Ligands and their Binding-Sites'''==
=== So4 ===
=== '''So4''' ===
In the gp12 there are two molecules of SO4 ligands. One of them <scene name='Sandbox_Reserved_717/Mynewscene/19'>SO4 (1529)</scene>  interacts with the citric acid. The blue molecule is the SO4. The other <scene name='Sandbox_Reserved_717/Mynewscene/21'>SO4 molecule (1530) interacts with Ser387</scene> . It builds two hydrogen bounds to Ser287 with the distances 2.66 and 3.20.  
In the gp12 there are two molecules of SO4 ligands. One of them <scene name='Sandbox_Reserved_717/Mynewscene/19'>SO4 (1529)</scene>  interacts with the citric acid. The blue molecule is the SO4. The other <scene name='Sandbox_Reserved_717/Mynewscene/21'>SO4 molecule (1530) interacts with Ser387</scene> . It builds two hydrogen bounds to Ser287 with the distances 2.66 and 3.20.  


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*[http://www.ebi.ac.uk/thornton-srv/databases/cgi-bin/pdbsum/GetPage.pl?pdbcode=1ocy&template=ligands.html&l=2.2]
*[http://www.ebi.ac.uk/thornton-srv/databases/cgi-bin/pdbsum/GetPage.pl?pdbcode=1ocy&template=ligands.html&l=2.2]


=== Citric Acid ===
=== '''Citric Acid''' ===
The <scene name='Sandbox_Reserved_717/Mynewscene/19'>citric acid </scene>. It is drawn in green. The citric acid interacts with following amino acids: Arg465 (distance 3.01), Asp455(distance 2.42) and Tyr454 (distance2.94). Here you can see the interactions:<scene name='Sandbox_Reserved_717/Mynewscene/22'>Interaction between citric acid, SO4, Asp465, Asp455 and Tyr454</scene>. These interactions are caused by hydrogen bounds.  
The <scene name='Sandbox_Reserved_717/Mynewscene/19'>citric acid </scene>. It is drawn in green. The citric acid interacts with following amino acids: Arg465 (distance 3.01), Asp455(distance 2.42) and Tyr454 (distance2.94). Here you can see the interactions:<scene name='Sandbox_Reserved_717/Mynewscene/22'>Interaction between citric acid, SO4, Asp465, Asp455 and Tyr454</scene>. These interactions are caused by hydrogen bounds.  


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*[http://www.ebi.ac.uk/thornton-srv/databases/cgi-bin/pdbsum/GetPage.pl?pdbcode=1ocy&template=ligands.html&l=1.1]
*[http://www.ebi.ac.uk/thornton-srv/databases/cgi-bin/pdbsum/GetPage.pl?pdbcode=1ocy&template=ligands.html&l=1.1]


=== Zinc ===
=== '''Zinc''' ===
The zinc is located in the centre of the receptor-binding domain. The zinc-binding site lies on the border between the head and the bonnet of the 45kDa domain. It interacts with the amino acids <scene name='Sandbox_Reserved_717/Mynewscene/3'>His445 and His447  </scene>  of each monomer. The distances of the zinc ion to the NE2 of <scene name='Sandbox_Reserved_717/Mynewscene/1'>His445</scene> and <scene name='Sandbox_Reserved_717/Mynewscene/2'>His447</scene> are 2.22 Å and 2.25 Å. The normally found distances between His and zinc are shorter. The explaination that these distances are longer than normally found is the octahedral coordination of the zinc in this structure.  
The zinc is located in the centre of the receptor-binding domain. The zinc-binding site lies on the border between the head and the bonnet of the 45kDa domain. It interacts with the amino acids <scene name='Sandbox_Reserved_717/Mynewscene/3'>His445 and His447  </scene>  of each monomer. The distances of the zinc ion to the NE2 of <scene name='Sandbox_Reserved_717/Mynewscene/1'>His445</scene> and <scene name='Sandbox_Reserved_717/Mynewscene/2'>His447</scene> are 2.22 Å and 2.25 Å. The normally found distances between His and zinc are shorter. The explaination that these distances are longer than normally found is the octahedral coordination of the zinc in this structure.  
The role of the zinc ion is probably absolute of structural nature. It increases the stability of the C-terminus of gp12 against proteases, but it also raises the stability of the C-terminus in general.
The role of the zinc ion is probably absolute of structural nature. It increases the stability of the C-terminus of gp12 against proteases, but it also raises the stability of the C-terminus in general.
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