Vanessa Chaplin/sandbox2 - Revision history

Vanessa Chaplin at 19:27, 3 December 2014

2014-12-03T19:27:06Z

← Older revision		Revision as of 19:27, 3 December 2014
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	The <scene name='40/400587/Active_site/8'>active site</scene>, which is located on a deep cleft between the β-strands comprising the DBSH core, contains the essential Fe(II). It is normally coordinated by the three Fe(II)-binding <scene name='40/400587/Active_site/14'>ligands</scene> formed by the conserved triad sequence, His-X-Asp/Glu-Xn-His.<ref name="review" /><ref>Schofield, C.J., Ratcliffe, P.J. "Signalling Bypoxia by HIF Hydroxylases." Biochemical and Biophysical Research Communications, August 24, 2005, 338, 617-626. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/16139242 16139242]</ref><ref name="structure" /> , 2-OG (which in this scene is replaced by HG) and a water molecule to form an octahedral geometry. Aside from the triad motif residues and those that bind 2-OG, the residues that are predominant inside the active site are nonpolar in nature. This is evidence of the enzyme's need to protect the protein core from oxidation by reactive species that are sometimes generated from iron-related reactions such as Fenton chemistry.<ref name="structure" />		The <scene name='40/400587/Active_site/8'>active site</scene>, which is located on a deep cleft between the β-strands comprising the DBSH core, contains the essential Fe(II). It is normally coordinated by the three Fe(II)-binding <scene name='40/400587/Active_site/14'>ligands</scene> formed by the conserved triad sequence, His-X-Asp/Glu-Xn-His.<ref name="review" /><ref>Schofield, C.J., Ratcliffe, P.J. "Signalling Bypoxia by HIF Hydroxylases." Biochemical and Biophysical Research Communications, August 24, 2005, 338, 617-626. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/16139242 16139242]</ref><ref name="structure" /> , 2-OG (which in this scene is replaced by HG) and a water molecule to form an octahedral geometry. Aside from the triad motif residues and those that bind 2-OG, the residues that are predominant inside the active site are nonpolar in nature. This is evidence of the enzyme's need to protect the protein core from oxidation by reactive species that are sometimes generated from iron-related reactions such as Fenton chemistry.<ref name="structure" />

	Upon binding of substrate, the <scene name='40/400587/B2b3_loop/2'>β2β3</scene> loop region of PHD2 undergoes a large <scene name='60/609795/B2b3_loop_with_cood/1'>conformational change</scene> ~~(Figure 1)~~. The β2β3 loop moves toward the active site and closes over the active site entrance. This loop movement is believed to be essential for the catalytic function of PHD2. The β2β3 loop region is mostly conserved in PHDs with minor differences. Mutational studies on the β2β3 loop region showed that this loop is significant for substrate recognition of PHDs<ref>PMID:18063574</ref>.		Upon binding of substrate, the <scene name='40/400587/B2b3_loop/2'>β2β3</scene> loop region of PHD2 (shown in purple) undergoes a large <scene name='60/609795/B2b3_loop_with_cood/1'>conformational change</scene>. The β2β3 loop moves toward the active site and closes over the active site entrance. This loop movement is believed to be essential for the catalytic function of PHD2. The β2β3 loop region is mostly conserved in PHDs with minor differences. Mutational studies on the β2β3 loop region showed that this loop is significant for substrate recognition of PHDs<ref>PMID:18063574</ref>.

	=== Function ===		=== Function ===

Vanessa Chaplin at 19:25, 3 December 2014

2014-12-03T19:25:12Z

← Older revision		Revision as of 19:25, 3 December 2014
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	<structure load='2g19' size='335' frame='true' align='right'		<structure load='2g19' size='335' frame='true' align='right'
	caption='Prolyl Hydroxylase Domain-2 (PHD-2) enzyme ~~with Fe+2 in complex with hydroxy-iodoisoquinolin-carbonyl glycine (HG)~~ [[2g19]]' />		caption='Significant structural features of Prolyl Hydroxylase Domain-2 (PHD-2) enzyme [[2g19]] and 3HQR' />

	<scene name='Molecular_Playground/Prolyl_Hydroxylase_Domain_(PHD)_Enzyme/Molecular_playground/4'>Molecular Playground</scene>		<scene name='Molecular_Playground/Prolyl_Hydroxylase_Domain_(PHD)_Enzyme/Molecular_playground/4'>Molecular Playground</scene>
Line 24:		Line 24:
	The <scene name='40/400587/Active_site/8'>active site</scene>, which is located on a deep cleft between the β-strands comprising the DBSH core, contains the essential Fe(II). It is normally coordinated by the three Fe(II)-binding <scene name='40/400587/Active_site/14'>ligands</scene> formed by the conserved triad sequence, His-X-Asp/Glu-Xn-His.<ref name="review" /><ref>Schofield, C.J., Ratcliffe, P.J. "Signalling Bypoxia by HIF Hydroxylases." Biochemical and Biophysical Research Communications, August 24, 2005, 338, 617-626. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/16139242 16139242]</ref><ref name="structure" /> , 2-OG (which in this scene is replaced by HG) and a water molecule to form an octahedral geometry. Aside from the triad motif residues and those that bind 2-OG, the residues that are predominant inside the active site are nonpolar in nature. This is evidence of the enzyme's need to protect the protein core from oxidation by reactive species that are sometimes generated from iron-related reactions such as Fenton chemistry.<ref name="structure" />		The <scene name='40/400587/Active_site/8'>active site</scene>, which is located on a deep cleft between the β-strands comprising the DBSH core, contains the essential Fe(II). It is normally coordinated by the three Fe(II)-binding <scene name='40/400587/Active_site/14'>ligands</scene> formed by the conserved triad sequence, His-X-Asp/Glu-Xn-His.<ref name="review" /><ref>Schofield, C.J., Ratcliffe, P.J. "Signalling Bypoxia by HIF Hydroxylases." Biochemical and Biophysical Research Communications, August 24, 2005, 338, 617-626. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/16139242 16139242]</ref><ref name="structure" /> , 2-OG (which in this scene is replaced by HG) and a water molecule to form an octahedral geometry. Aside from the triad motif residues and those that bind 2-OG, the residues that are predominant inside the active site are nonpolar in nature. This is evidence of the enzyme's need to protect the protein core from oxidation by reactive species that are sometimes generated from iron-related reactions such as Fenton chemistry.<ref name="structure" />

	Upon binding of substrate, the <scene name='40/400587/B2b3_loop/2'>β2β3</scene> loop region of PHD2 undergoes a large <scene name='60/609795/B2b3_loop_with_cood/1'>conformational change</scene> (Figure 1). The β2β3 loop moves toward the active site and closes over the active site entrance. This loop movement is believed to be essential for the catalytic function of PHD2. The β2β3 loop region is mostly conserved in PHDs with minor differences. Mutational studies on the β2β3 loop region showed that this loop is significant for substrate recognition of PHDs<ref>PMID:18063574</ref>.		Upon binding of substrate, the <scene name='40/400587/B2b3_loop/2'>β2β3</scene> loop region of PHD2 undergoes a large <scene name='60/609795/B2b3_loop_with_cood/1'>conformational change</scene> (Figure 1). The β2β3 loop moves toward the active site and closes over the active site entrance. This loop movement is believed to be essential for the catalytic function of PHD2. The β2β3 loop region is mostly conserved in PHDs with minor differences. Mutational studies on the β2β3 loop region showed that this loop is significant for substrate recognition of PHDs<ref>PMID:18063574</ref>.

	~~[[Image:PHD2.png\|315 px\|thumb\|Fig. 1: Substrate bound (purple) and unbound (green) PHD2. Figure created with PyMol. PDB IDs 2G19, 3HQR.]]~~

	=== Function ===		=== Function ===

Vanessa Chaplin at 19:21, 3 December 2014

2014-12-03T19:21:21Z

← Older revision		Revision as of 19:21, 3 December 2014
Line 24:		Line 24:
	The <scene name='40/400587/Active_site/8'>active site</scene>, which is located on a deep cleft between the β-strands comprising the DBSH core, contains the essential Fe(II). It is normally coordinated by the three Fe(II)-binding <scene name='40/400587/Active_site/14'>ligands</scene> formed by the conserved triad sequence, His-X-Asp/Glu-Xn-His.<ref name="review" /><ref>Schofield, C.J., Ratcliffe, P.J. "Signalling Bypoxia by HIF Hydroxylases." Biochemical and Biophysical Research Communications, August 24, 2005, 338, 617-626. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/16139242 16139242]</ref><ref name="structure" /> , 2-OG (which in this scene is replaced by HG) and a water molecule to form an octahedral geometry. Aside from the triad motif residues and those that bind 2-OG, the residues that are predominant inside the active site are nonpolar in nature. This is evidence of the enzyme's need to protect the protein core from oxidation by reactive species that are sometimes generated from iron-related reactions such as Fenton chemistry.<ref name="structure" />		The <scene name='40/400587/Active_site/8'>active site</scene>, which is located on a deep cleft between the β-strands comprising the DBSH core, contains the essential Fe(II). It is normally coordinated by the three Fe(II)-binding <scene name='40/400587/Active_site/14'>ligands</scene> formed by the conserved triad sequence, His-X-Asp/Glu-Xn-His.<ref name="review" /><ref>Schofield, C.J., Ratcliffe, P.J. "Signalling Bypoxia by HIF Hydroxylases." Biochemical and Biophysical Research Communications, August 24, 2005, 338, 617-626. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/16139242 16139242]</ref><ref name="structure" /> , 2-OG (which in this scene is replaced by HG) and a water molecule to form an octahedral geometry. Aside from the triad motif residues and those that bind 2-OG, the residues that are predominant inside the active site are nonpolar in nature. This is evidence of the enzyme's need to protect the protein core from oxidation by reactive species that are sometimes generated from iron-related reactions such as Fenton chemistry.<ref name="structure" />

	Upon binding of substrate, the <scene name='40/400587/B2b3_loop/2'>β2β3</scene> loop region of PHD2 undergoes a large conformational change (Figure 1). The β2β3 loop moves toward the active site and closes over the active site entrance. This loop movement is believed to be essential for the catalytic function of PHD2. The β2β3 loop region is mostly conserved in PHDs with minor differences. Mutational studies on the β2β3 loop region showed that this loop is significant for substrate recognition of PHDs<ref>PMID:18063574</ref>.		Upon binding of substrate, the <scene name='40/400587/B2b3_loop/2'>β2β3</scene> loop region of PHD2 undergoes a large <scene name='60/609795/B2b3_loop_with_cood/1'>conformational change</scene> (Figure 1). The β2β3 loop moves toward the active site and closes over the active site entrance. This loop movement is believed to be essential for the catalytic function of PHD2. The β2β3 loop region is mostly conserved in PHDs with minor differences. Mutational studies on the β2β3 loop region showed that this loop is significant for substrate recognition of PHDs<ref>PMID:18063574</ref>.

	[[Image:PHD2.png\|315 px\|thumb\|Fig. 1: Substrate bound (purple) and unbound (green) PHD2. Figure created with PyMol. PDB IDs 2G19, 3HQR.]]		[[Image:PHD2.png\|315 px\|thumb\|Fig. 1: Substrate bound (purple) and unbound (green) PHD2. Figure created with PyMol. PDB IDs 2G19, 3HQR.]]

Vanessa Chaplin at 20:41, 21 November 2014

2014-11-21T20:41:50Z

Vanessa Chaplin at 20:21, 21 November 2014

2014-11-21T20:21:20Z

← Older revision		Revision as of 20:21, 21 November 2014
Line 6:		Line 6:
	Metazoans adapt to oxygen levels in the environment by making use of intracellular oxygen levels as signals to regulate the [http://en.wikipedia.org/wiki/Transcription_(genetics) transcription] of genes that are essential under normoxic or [http://en.wikipedia.org/wiki/Hypoxia_(medical) hypoxic] conditions. Central to this mechanism is the oxygen-dependent hydroxylation of specific proline and asparagine residues of the transcription factor, hypoxia-inducible factor [http://en.wikipedia.org/wiki/HIF1A (HIF)-α] by the HIF-hydroxylase enzymes, which are studied extensively by the [http://people.chem.umass.edu/knapplab/?q=knappchem/index.html Knapp Lab] here at UMass Amherst.<ref name="review">Fong, G.H., Takeda, K. "Role and Regulation of Prolyl Hydroxylase Domain Proteins." Cell Death and Differentiation, February 15, 2008, 15, 635-641. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/18259202 18259202]</ref>		Metazoans adapt to oxygen levels in the environment by making use of intracellular oxygen levels as signals to regulate the [http://en.wikipedia.org/wiki/Transcription_(genetics) transcription] of genes that are essential under normoxic or [http://en.wikipedia.org/wiki/Hypoxia_(medical) hypoxic] conditions. Central to this mechanism is the oxygen-dependent hydroxylation of specific proline and asparagine residues of the transcription factor, hypoxia-inducible factor [http://en.wikipedia.org/wiki/HIF1A (HIF)-α] by the HIF-hydroxylase enzymes, which are studied extensively by the [http://people.chem.umass.edu/knapplab/?q=knappchem/index.html Knapp Lab] here at UMass Amherst.<ref name="review">Fong, G.H., Takeda, K. "Role and Regulation of Prolyl Hydroxylase Domain Proteins." Cell Death and Differentiation, February 15, 2008, 15, 635-641. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/18259202 18259202]</ref>

	These HIF-hydroxylase enzymes include Prolyl Hydroxylase Domain isoforms (PHDs) and Factor Inhibition Hypoxia Inducible Factor (FIH). PHDs belong to the same oxygenase superfamily as the [http://en.wikipedia.org/wiki/Collagen collagen] prolyl hydroxylases. Inside the cell, PHD1 is found in the nucleus, while PHD2 is mostly located in the cytoplasm and PHD3 is distributed in both <ref>PMID:12615973</ref>.This distribution contrasts with that of the [http://en.wikipedia.org/wiki/Collagen collagen] prolyl hydroxylases, which reside in the endoplasmic reticulum. In mammals, the PHD dioxygenase subfamily originally included three [http://en.wiktionary.org/wiki/homolog homolog] members: PHD1 (also known as HPH3 and [http://en.wikipedia.org/wiki/EGLN2 EGLN2]), PHD2 (also known as HPH2 and [http://en.wikipedia.org/wiki/EGLN1 EGLN1]), PHD3 (also known as HPH1 and [http://en.wikipedia.org/wiki/EGLN3 EGLN3]). Recently, a newly identified enzyme called P4H-TM (also named PHD4 and EGLN4) was added to this subfamily. Both PHD1 and PHD2 contain more than 400 [http://en.wikipedia.org/wiki/Amino_acid amino acid] residues while PHD3 has less than 250. All isoforms, however, contain the highly conserved hydroxylase domain in the catalytic carboxy-terminal region. <ref name="review" />. PHDs differ in terms of the proline hydroxylation site. PHD2 hydroxylates Pro-402 or Pro-564 ~~residues~~ in the oxygen degradation domain of [http://en.wikipedia.org/wiki/HIF1A (HIF)-α] whereas PHD3 can only hydroxylate the Pro-564 position.		These HIF-hydroxylase enzymes include the Prolyl Hydroxylase Domain isoforms (PHDs) and Factor Inhibition Hypoxia Inducible Factor (FIH). PHDs belong to the same oxygenase superfamily as the [http://en.wikipedia.org/wiki/Collagen collagen] prolyl hydroxylases. Inside the cell, PHD1 is found in the nucleus, while PHD2 is mostly located in the cytoplasm and PHD3 is distributed in both. <ref>PMID:12615973</ref> This distribution contrasts with that of the [http://en.wikipedia.org/wiki/Collagen collagen] prolyl hydroxylases, which reside in the endoplasmic reticulum. In mammals, the PHD dioxygenase subfamily originally included three [http://en.wiktionary.org/wiki/homolog homolog] members: PHD1 (also known as HPH3 and [http://en.wikipedia.org/wiki/EGLN2 EGLN2]), PHD2 (also known as HPH2 and [http://en.wikipedia.org/wiki/EGLN1 EGLN1]), PHD3 (also known as HPH1 and [http://en.wikipedia.org/wiki/EGLN3 EGLN3]). Recently, a newly identified enzyme called P4H-TM (also named PHD4 and EGLN4) was added to this subfamily. Both PHD1 and PHD2 contain more than 400 [http://en.wikipedia.org/wiki/Amino_acid amino acid] residues while PHD3 has less than 250. All isoforms, however, contain the highly conserved hydroxylase domain in the catalytic carboxy-terminal region. <ref name="review" /> PHDs differ in terms of the proline hydroxylation site. PHD2 hydroxylates either the Pro-402 or Pro-564 residue in the oxygen degradation domain of [http://en.wikipedia.org/wiki/HIF1A (HIF)-α] whereas PHD3 can only hydroxylate the Pro-564 position.

	Of the HIF-hydroxylases, '''[http://www.chem.qmul.ac.uk/iubmb/enzyme/EC1/14/11/ (PHD2)]''' is believed to be the main oxygen sensor. PHD2 is a Fe(II)/2-oxoglutarate (OG)-dependent [http://en.wikipedia.org/wiki/Oxygenase dioxygenase] that catalyzes the ''trans''-4-hydroxylation of specific proline residues (in humans, Pro-402 and Pro-564) in [http://en.wikipedia.org/wiki/HIF1A (HIF)-α]. In addition to iron, this enzyme also requires [http://en.wikipedia.org/wiki/Vitamin_C ascorbate] as a cofactor.<ref name="structure">Mcdonough, M.A., Li, V., Flashman, E., et al. "Cellular oxygen sensing: Crystal structure of hypoxia-inducible factor prolyl hydroxylase (PHD2)." PNAS, June 27, 2006, 103 (26), 9814-9819. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/16782814 16782814]</ref>

			Of the HIF-hydroxylases, '''[http://www.chem.qmul.ac.uk/iubmb/enzyme/EC1/14/11/ (PHD2)]''' is believed to be the main oxygen sensor. PHD2 is a Fe(II)/2-oxoglutarate (OG)-dependent [http://en.wikipedia.org/wiki/Oxygenase dioxygenase] that catalyzes the ''trans''-4-hydroxylation of specific proline residues (as mentioned above) in [http://en.wikipedia.org/wiki/HIF1A (HIF)-α]. In addition to iron, this enzyme also requires [http://en.wikipedia.org/wiki/Vitamin_C ascorbate] as a cofactor.<ref name="structure">Mcdonough, M.A., Li, V., Flashman, E., et al. "Cellular oxygen sensing: Crystal structure of hypoxia-inducible factor prolyl hydroxylase (PHD2)." PNAS, June 27, 2006, 103 (26), 9814-9819. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/16782814 16782814]</ref> Our goal in the Knapp Lab is to understand and define the mechanism of action for this enzyme and evaluate how oxygen affects enzyme activity to gaining further knowledge of oxygen sensing.

	<structure load='2g19' size='335' frame='true' align='right'		<structure load='2g19' size='335' frame='true' align='right'
	caption='Prolyl Hydroxylase Domain-2 (PHD-2) enzyme with Fe+2 in complex with hydroxy-iodoisoquinolin-carbonyl glycine (HG) [[2g19]]' />		caption='Prolyl Hydroxylase Domain-2 (PHD-2) enzyme with Fe+2 in complex with hydroxy-iodoisoquinolin-carbonyl glycine (HG) [[2g19]]' />


	<scene name='Molecular_Playground/Prolyl_Hydroxylase_Domain_(PHD)_Enzyme/Molecular_playground/4'>Molecular Playground</scene>		<scene name='Molecular_Playground/Prolyl_Hydroxylase_Domain_(PHD)_Enzyme/Molecular_playground/4'>Molecular Playground</scene>

Vanessa Chaplin at 19:45, 21 November 2014

2014-11-21T19:45:23Z

← Older revision		Revision as of 19:45, 21 November 2014
Line 6:		Line 6:
	Metazoans adapt to oxygen levels in the environment by making use of intracellular oxygen levels as signals to regulate the [http://en.wikipedia.org/wiki/Transcription_(genetics) transcription] of genes that are essential under normoxic or [http://en.wikipedia.org/wiki/Hypoxia_(medical) hypoxic] conditions. Central to this mechanism is the oxygen-dependent hydroxylation of specific proline and asparagine residues of the transcription factor, hypoxia-inducible factor [http://en.wikipedia.org/wiki/HIF1A (HIF)-α] by the HIF-hydroxylase enzymes, which are studied extensively by the [http://people.chem.umass.edu/knapplab/?q=knappchem/index.html Knapp Lab] here at UMass Amherst.<ref name="review">Fong, G.H., Takeda, K. "Role and Regulation of Prolyl Hydroxylase Domain Proteins." Cell Death and Differentiation, February 15, 2008, 15, 635-641. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/18259202 18259202]</ref>		Metazoans adapt to oxygen levels in the environment by making use of intracellular oxygen levels as signals to regulate the [http://en.wikipedia.org/wiki/Transcription_(genetics) transcription] of genes that are essential under normoxic or [http://en.wikipedia.org/wiki/Hypoxia_(medical) hypoxic] conditions. Central to this mechanism is the oxygen-dependent hydroxylation of specific proline and asparagine residues of the transcription factor, hypoxia-inducible factor [http://en.wikipedia.org/wiki/HIF1A (HIF)-α] by the HIF-hydroxylase enzymes, which are studied extensively by the [http://people.chem.umass.edu/knapplab/?q=knappchem/index.html Knapp Lab] here at UMass Amherst.<ref name="review">Fong, G.H., Takeda, K. "Role and Regulation of Prolyl Hydroxylase Domain Proteins." Cell Death and Differentiation, February 15, 2008, 15, 635-641. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/18259202 18259202]</ref>

	Of the HIF-hydroxylases, '''~~Prolyl hydroxylase domain~~ [http://www.chem.qmul.ac.uk/iubmb/enzyme/EC1/14/11/ (~~PHD~~)]''' is believed to be the main oxygen sensor. PHD2 is a Fe(II)/2-oxoglutarate (OG)-dependent [http://en.wikipedia.org/wiki/Oxygenase dioxygenase] that catalyzes the ''trans''-4-hydroxylation of specific proline residues (in humans, Pro-402 and Pro-564) in [http://en.wikipedia.org/wiki/HIF1A (HIF)-α]. In addition to iron, this enzyme also requires [http://en.wikipedia.org/wiki/Vitamin_C ascorbate] as a cofactor.<ref name="structure">Mcdonough, M.A., Li, V., Flashman, E., et al. "Cellular oxygen sensing: Crystal structure of hypoxia-inducible factor prolyl hydroxylase (PHD2)." PNAS, June 27, 2006, 103 (26), 9814-9819. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/16782814 16782814]</ref>		These HIF-hydroxylase enzymes include Prolyl Hydroxylase Domain isoforms (PHDs) and Factor Inhibition Hypoxia Inducible Factor (FIH). PHDs belong to the same oxygenase superfamily as the [http://en.wikipedia.org/wiki/Collagen collagen] prolyl hydroxylases. Inside the cell, PHD1 is found in the nucleus, while PHD2 is mostly located in the cytoplasm and PHD3 is distributed in both <ref>PMID:12615973</ref>.This distribution contrasts with that of the [http://en.wikipedia.org/wiki/Collagen collagen] prolyl hydroxylases, which reside in the endoplasmic reticulum. In mammals, the PHD dioxygenase subfamily originally included three [http://en.wiktionary.org/wiki/homolog homolog] members: PHD1 (also known as HPH3 and [http://en.wikipedia.org/wiki/EGLN2 EGLN2]), PHD2 (also known as HPH2 and [http://en.wikipedia.org/wiki/EGLN1 EGLN1]), PHD3 (also known as HPH1 and [http://en.wikipedia.org/wiki/EGLN3 EGLN3]). Recently, a newly identified enzyme called P4H-TM (also named PHD4 and EGLN4) was added to this subfamily. Both PHD1 and PHD2 contain more than 400 [http://en.wikipedia.org/wiki/Amino_acid amino acid] residues while PHD3 has less than 250. All isoforms, however, contain the highly conserved hydroxylase domain in the catalytic carboxy-terminal region. <ref name="review" />. PHDs differ in terms of the proline hydroxylation site. PHD2 hydroxylates Pro-402 or Pro-564 residues in the oxygen degradation domain of [http://en.wikipedia.org/wiki/HIF1A (HIF)-α] whereas PHD3 can only hydroxylate the Pro-564 position.

			Of the HIF-hydroxylases, '''[http://www.chem.qmul.ac.uk/iubmb/enzyme/EC1/14/11/ (PHD2)]''' is believed to be the main oxygen sensor. PHD2 is a Fe(II)/2-oxoglutarate (OG)-dependent [http://en.wikipedia.org/wiki/Oxygenase dioxygenase] that catalyzes the ''trans''-4-hydroxylation of specific proline residues (in humans, Pro-402 and Pro-564) in [http://en.wikipedia.org/wiki/HIF1A (HIF)-α]. In addition to iron, this enzyme also requires [http://en.wikipedia.org/wiki/Vitamin_C ascorbate] as a cofactor.<ref name="structure">Mcdonough, M.A., Li, V., Flashman, E., et al. "Cellular oxygen sensing: Crystal structure of hypoxia-inducible factor prolyl hydroxylase (PHD2)." PNAS, June 27, 2006, 103 (26), 9814-9819. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/16782814 16782814]</ref>

	PHDs belong to the same oxygenase superfamily as the [http://en.wikipedia.org/wiki/Collagen collagen] prolyl hydroxylases. Inside the cell, PHD1 is found in the nucleus, PHD2 is mostly located in the cytoplasm and PHD3 is distributed in both <ref>PMID:12615973</ref> in contrast to [http://en.wikipedia.org/wiki/Collagen collagen] prolyl hydroxylases, which reside in the endoplasmic reticulum. In mammals, the PHD dioxygenase subfamily originally includes three [http://en.wiktionary.org/wiki/homolog homolog] members but was recently updated to include another member: PHD1 (also known as HPH3 and [http://en.wikipedia.org/wiki/EGLN2 EGLN2]), PHD2 (also known as HPH2 and [http://en.wikipedia.org/wiki/EGLN1 EGLN1]), PHD3 (also known as HPH1 and [http://en.wikipedia.org/wiki/EGLN3 EGLN3]), and a newly identified enzyme called P4H-TM (also recently named PHD4 and EGLN4). Both PHD1 and PHD2 contain more than 400 [http://en.wikipedia.org/wiki/Amino_acid amino acid] residues while PHD3 has less than 250. All isoforms, however, contain the highly conserved hydroxylase domain in the catalytic carboxy-terminal region. <ref name="review" />. PHDs differ in terms of the proline hydroxylation site. PHD2 hydroxylates Pro-402 and Pro-564 residues in the oxygen degradation domain of [http://en.wikipedia.org/wiki/HIF1A (HIF)-α] whereas PHD3 can only hydroxylate Pro-564 position.

	<structure load='2g19' size='335' frame='true' align='right'		<structure load='2g19' size='335' frame='true' align='right'

Vanessa Chaplin at 19:35, 21 November 2014

2014-11-21T19:35:47Z

← Older revision		Revision as of 19:35, 21 November 2014
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	Metazoans adapt to oxygen levels in the environment by making use of intracellular oxygen levels as signals to regulate the [http://en.wikipedia.org/wiki/Transcription_(genetics) transcription] of genes that are essential under normoxic or [http://en.wikipedia.org/wiki/Hypoxia_(medical) hypoxic] conditions. Central to this mechanism is the oxygen-dependent hydroxylation of specific proline and asparagine residues of the transcription factor, hypoxia-inducible factor [http://en.wikipedia.org/wiki/HIF1A (HIF)-α] by the HIF-hydroxylase enzymes, which are studied extensively by the [http://people.chem.umass.edu/knapplab/?q=knappchem/index.html Knapp Lab] here at UMass Amherst.<ref name="review">Fong, G.H., Takeda, K. "Role and Regulation of Prolyl Hydroxylase Domain Proteins." Cell Death and Differentiation, February 15, 2008, 15, 635-641. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/18259202 18259202]</ref>		Metazoans adapt to oxygen levels in the environment by making use of intracellular oxygen levels as signals to regulate the [http://en.wikipedia.org/wiki/Transcription_(genetics) transcription] of genes that are essential under normoxic or [http://en.wikipedia.org/wiki/Hypoxia_(medical) hypoxic] conditions. Central to this mechanism is the oxygen-dependent hydroxylation of specific proline and asparagine residues of the transcription factor, hypoxia-inducible factor [http://en.wikipedia.org/wiki/HIF1A (HIF)-α] by the HIF-hydroxylase enzymes, which are studied extensively by the [http://people.chem.umass.edu/knapplab/?q=knappchem/index.html Knapp Lab] here at UMass Amherst.<ref name="review">Fong, G.H., Takeda, K. "Role and Regulation of Prolyl Hydroxylase Domain Proteins." Cell Death and Differentiation, February 15, 2008, 15, 635-641. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/18259202 18259202]</ref>

	'''Prolyl hydroxylase domain ~~(PHD) enzyme'''~~ [http://www.chem.qmul.ac.uk/iubmb/enzyme/EC1/14/11/ (~~EC 1.14.11.-~~)] is a Fe(II)/2-oxoglutarate (OG)-dependent [http://en.wikipedia.org/wiki/Oxygenase dioxygenase] that catalyzes the ''trans''-4-hydroxylation of specific proline residues (in humans, Pro-402 and Pro-564) in [http://en.wikipedia.org/wiki/HIF1A (HIF)-α]. In addition to iron, this enzyme also requires [http://en.wikipedia.org/wiki/Vitamin_C ascorbate] as a cofactor.<ref name="structure">Mcdonough, M.A., Li, V., Flashman, E., et al. "Cellular oxygen sensing: Crystal structure of hypoxia-inducible factor prolyl hydroxylase (PHD2)." PNAS, June 27, 2006, 103 (26), 9814-9819. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/16782814 16782814]</ref>		Of the HIF-hydroxylases, '''Prolyl hydroxylase domain [http://www.chem.qmul.ac.uk/iubmb/enzyme/EC1/14/11/ (PHD)]''' is believed to be the main oxygen sensor. PHD2 is a Fe(II)/2-oxoglutarate (OG)-dependent [http://en.wikipedia.org/wiki/Oxygenase dioxygenase] that catalyzes the ''trans''-4-hydroxylation of specific proline residues (in humans, Pro-402 and Pro-564) in [http://en.wikipedia.org/wiki/HIF1A (HIF)-α]. In addition to iron, this enzyme also requires [http://en.wikipedia.org/wiki/Vitamin_C ascorbate] as a cofactor.<ref name="structure">Mcdonough, M.A., Li, V., Flashman, E., et al. "Cellular oxygen sensing: Crystal structure of hypoxia-inducible factor prolyl hydroxylase (PHD2)." PNAS, June 27, 2006, 103 (26), 9814-9819. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/16782814 16782814]</ref>

	PHDs belong to the same oxygenase superfamily as the [http://en.wikipedia.org/wiki/Collagen collagen] prolyl hydroxylases. Inside the cell, PHD1 is found in the nucleus, PHD2 is mostly located in the cytoplasm and PHD3 is distributed in both <ref>PMID:12615973</ref> in contrast to [http://en.wikipedia.org/wiki/Collagen collagen] prolyl hydroxylases, which reside in the endoplasmic reticulum. In mammals, the PHD dioxygenase subfamily originally includes three [http://en.wiktionary.org/wiki/homolog homolog] members but was recently updated to include another member: PHD1 (also known as HPH3 and [http://en.wikipedia.org/wiki/EGLN2 EGLN2]), PHD2 (also known as HPH2 and [http://en.wikipedia.org/wiki/EGLN1 EGLN1]), PHD3 (also known as HPH1 and [http://en.wikipedia.org/wiki/EGLN3 EGLN3]), and a newly identified enzyme called P4H-TM (also recently named PHD4 and EGLN4). Both PHD1 and PHD2 contain more than 400 [http://en.wikipedia.org/wiki/Amino_acid amino acid] residues while PHD3 has less than 250. All isoforms, however, contain the highly conserved hydroxylase domain in the catalytic carboxy-terminal region. <ref name="review" />. PHDs differ in terms of the proline hydroxylation site. PHD2 hydroxylates Pro-402 and Pro-564 residues in the oxygen degradation domain of [http://en.wikipedia.org/wiki/HIF1A (HIF)-α] whereas PHD3 can only hydroxylate Pro-564 position.		PHDs belong to the same oxygenase superfamily as the [http://en.wikipedia.org/wiki/Collagen collagen] prolyl hydroxylases. Inside the cell, PHD1 is found in the nucleus, PHD2 is mostly located in the cytoplasm and PHD3 is distributed in both <ref>PMID:12615973</ref> in contrast to [http://en.wikipedia.org/wiki/Collagen collagen] prolyl hydroxylases, which reside in the endoplasmic reticulum. In mammals, the PHD dioxygenase subfamily originally includes three [http://en.wiktionary.org/wiki/homolog homolog] members but was recently updated to include another member: PHD1 (also known as HPH3 and [http://en.wikipedia.org/wiki/EGLN2 EGLN2]), PHD2 (also known as HPH2 and [http://en.wikipedia.org/wiki/EGLN1 EGLN1]), PHD3 (also known as HPH1 and [http://en.wikipedia.org/wiki/EGLN3 EGLN3]), and a newly identified enzyme called P4H-TM (also recently named PHD4 and EGLN4). Both PHD1 and PHD2 contain more than 400 [http://en.wikipedia.org/wiki/Amino_acid amino acid] residues while PHD3 has less than 250. All isoforms, however, contain the highly conserved hydroxylase domain in the catalytic carboxy-terminal region. <ref name="review" />. PHDs differ in terms of the proline hydroxylation site. PHD2 hydroxylates Pro-402 and Pro-564 residues in the oxygen degradation domain of [http://en.wikipedia.org/wiki/HIF1A (HIF)-α] whereas PHD3 can only hydroxylate Pro-564 position.

Vanessa Chaplin at 19:32, 21 November 2014

2014-11-21T19:32:07Z

← Older revision		Revision as of 19:32, 21 November 2014
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	=== Introduction ===		=== Introduction ===

	Metazoans adapt to oxygen levels in the environment by making use of intracellular oxygen levels as signals to regulate the [http://en.wikipedia.org/wiki/Transcription_(genetics) transcription] of genes that are essential under normoxic or [http://en.wikipedia.org/wiki/Hypoxia_(medical) hypoxic] conditions. Central to this mechanism is the oxygen-dependent hydroxylation of specific proline and asparagine residues of the transcription factor, hypoxia-inducible factor [http://en.wikipedia.org/wiki/HIF1A (HIF)-α].<ref name="review">Fong, G.H., Takeda, K. "Role and Regulation of Prolyl Hydroxylase Domain Proteins." Cell Death and Differentiation, February 15, 2008, 15, 635-641. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/18259202 18259202]</ref>		Metazoans adapt to oxygen levels in the environment by making use of intracellular oxygen levels as signals to regulate the [http://en.wikipedia.org/wiki/Transcription_(genetics) transcription] of genes that are essential under normoxic or [http://en.wikipedia.org/wiki/Hypoxia_(medical) hypoxic] conditions. Central to this mechanism is the oxygen-dependent hydroxylation of specific proline and asparagine residues of the transcription factor, hypoxia-inducible factor [http://en.wikipedia.org/wiki/HIF1A (HIF)-α] by the HIF-hydroxylase enzymes, which are studied extensively by the [http://people.chem.umass.edu/knapplab/?q=knappchem/index.html Knapp Lab] here at UMass Amherst.<ref name="review">Fong, G.H., Takeda, K. "Role and Regulation of Prolyl Hydroxylase Domain Proteins." Cell Death and Differentiation, February 15, 2008, 15, 635-641. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/18259202 18259202]</ref>

	'''Prolyl hydroxylase domain (PHD) enzyme''' [http://www.chem.qmul.ac.uk/iubmb/enzyme/EC1/14/11/ (EC 1.14.11.-)] is a Fe(II)/2-oxoglutarate (OG)-dependent [http://en.wikipedia.org/wiki/Oxygenase dioxygenase] that catalyzes the ''trans''-4-hydroxylation of specific proline residues (in humans, Pro-402 and Pro-564) in [http://en.wikipedia.org/wiki/HIF1A (HIF)-α]. In addition to iron, this enzyme also requires [http://en.wikipedia.org/wiki/Vitamin_C ascorbate] as a cofactor.<ref name="structure">Mcdonough, M.A., Li, V., Flashman, E., et al. "Cellular oxygen sensing: Crystal structure of hypoxia-inducible factor prolyl hydroxylase (PHD2)." PNAS, June 27, 2006, 103 (26), 9814-9819. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/16782814 16782814]</ref>		'''Prolyl hydroxylase domain (PHD) enzyme''' [http://www.chem.qmul.ac.uk/iubmb/enzyme/EC1/14/11/ (EC 1.14.11.-)] is a Fe(II)/2-oxoglutarate (OG)-dependent [http://en.wikipedia.org/wiki/Oxygenase dioxygenase] that catalyzes the ''trans''-4-hydroxylation of specific proline residues (in humans, Pro-402 and Pro-564) in [http://en.wikipedia.org/wiki/HIF1A (HIF)-α]. In addition to iron, this enzyme also requires [http://en.wikipedia.org/wiki/Vitamin_C ascorbate] as a cofactor.<ref name="structure">Mcdonough, M.A., Li, V., Flashman, E., et al. "Cellular oxygen sensing: Crystal structure of hypoxia-inducible factor prolyl hydroxylase (PHD2)." PNAS, June 27, 2006, 103 (26), 9814-9819. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/16782814 16782814]</ref>

Vanessa Chaplin at 18:01, 19 November 2014

2014-11-19T18:01:36Z

← Older revision		Revision as of 18:01, 19 November 2014
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	~~==Your Heading Here (maybe something like 'Structure')==~~		One of the [[CBI Molecules]] being studied in the [http://www.umass.edu/cbi/ University of Massachusetts Amherst Chemistry-Biology Interface Program] at UMass Amherst and on display at the [http://www.molecularplayground.org/ Molecular Playground].
	~~<StructureSection load='1stp' size='340' side='right' caption='Caption for this structure' scene=''>~~
	~~This is a default text for your page '''Vanessa Chaplin/sandbox2'''. Click above on '''edit this page''' to modify. Be careful with~~ the ~~< and > signs.~~
	~~You may include any references to papers as~~ in: ~~the use of JSmol in Proteopedia <ref>DOI 10~~.~~1002~~/~~ijch.201300024<~~/~~ref> or to~~ the ~~article describing Jmol <ref>PMID~~:~~21638687<~~/~~ref> to the rescue~~.

	~~== Function ==~~

	== ~~Disease~~ ==		=== Introduction ===

	=~~= Relevance ==~~		Metazoans adapt to oxygen levels in the environment by making use of intracellular oxygen levels as signals to regulate the [http://en.wikipedia.org/wiki/Transcription_(genetics) transcription] of genes that are essential under normoxic or [http://en.wikipedia.org/wiki/Hypoxia_(medical) hypoxic] conditions. Central to this mechanism is the oxygen-dependent hydroxylation of specific proline and asparagine residues of the transcription factor, hypoxia-inducible factor [http://en.wikipedia.org/wiki/HIF1A (HIF)-α].<ref name="review">Fong, G.H., Takeda, K. "Role and Regulation of Prolyl Hydroxylase Domain Proteins." Cell Death and Differentiation, February 15, 2008, 15, 635-641. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/18259202 18259202]</ref>

	=~~= Structural highlights ==~~		'''Prolyl hydroxylase domain (PHD) enzyme''' [http://www.chem.qmul.ac.uk/iubmb/enzyme/EC1/14/11/ (EC 1.14.11.-)] is a Fe(II)/2-oxoglutarate (OG)-dependent [http://en.wikipedia.org/wiki/Oxygenase dioxygenase] that catalyzes the ''trans''-4-hydroxylation of specific proline residues (in humans, Pro-402 and Pro-564) in [http://en.wikipedia.org/wiki/HIF1A (HIF)-α]. In addition to iron, this enzyme also requires [http://en.wikipedia.org/wiki/Vitamin_C ascorbate] as a cofactor.<ref name="structure">Mcdonough, M.A., Li, V., Flashman, E., et al. "Cellular oxygen sensing: Crystal structure of hypoxia-inducible factor prolyl hydroxylase (PHD2)." PNAS, June 27, 2006, 103 (26), 9814-9819. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/16782814 16782814]</ref>

			PHDs belong to the same oxygenase superfamily as the [http://en.wikipedia.org/wiki/Collagen collagen] prolyl hydroxylases. Inside the cell, PHD1 is found in the nucleus, PHD2 is mostly located in the cytoplasm and PHD3 is distributed in both <ref>PMID:12615973</ref> in contrast to [http://en.wikipedia.org/wiki/Collagen collagen] prolyl hydroxylases, which reside in the endoplasmic reticulum. In mammals, the PHD dioxygenase subfamily originally includes three [http://en.wiktionary.org/wiki/homolog homolog] members but was recently updated to include another member: PHD1 (also known as HPH3 and [http://en.wikipedia.org/wiki/EGLN2 EGLN2]), PHD2 (also known as HPH2 and [http://en.wikipedia.org/wiki/EGLN1 EGLN1]), PHD3 (also known as HPH1 and [http://en.wikipedia.org/wiki/EGLN3 EGLN3]), and a newly identified enzyme called P4H-TM (also recently named PHD4 and EGLN4). Both PHD1 and PHD2 contain more than 400 [http://en.wikipedia.org/wiki/Amino_acid amino acid] residues while PHD3 has less than 250. All isoforms, however, contain the highly conserved hydroxylase domain in the catalytic carboxy-terminal region. <ref name="review" />. PHDs differ in terms of the proline hydroxylation site. PHD2 hydroxylates Pro-402 and Pro-564 residues in the oxygen degradation domain of [http://en.wikipedia.org/wiki/HIF1A (HIF)-α] whereas PHD3 can only hydroxylate Pro-564 position.

	== ~~Headline text~~ ==		<structure load='2g19' size='335' frame='true' align='right'
			caption='Prolyl Hydroxylase Domain-2 (PHD-2) enzyme with Fe+2 in complex with hydroxy-iodoisoquinolin-carbonyl glycine (HG) [[2g19]]' />


	~~This is a sample scene created with SAT to~~ <scene name=~~"/12/3456/Sample/1">color</scene> by Group, and another to make <scene name="/12~~/~~3456~~/~~Sample~~/2">~~a transparent representation~~</scene> ~~of the protein. You can make your own scenes on SAT starting from scratch or loading and editing one of these sample scenes.~~		<scene name='Molecular_Playground/Prolyl_Hydroxylase_Domain_(PHD)_Enzyme/Molecular_playground/4'>Molecular Playground</scene>

	</~~StructureSection~~>		Molecular Playground banner: Prolyl Hydroxylase Domain (PHD) enzyme, a cellular oxygen sensor, has a major regulatory role in oxygen homeostasis.
	== References ==
			=== Structure ===
			----

			PHDs have two structural domains: the more variable N-terminal domain and the conserved catalytic C-terminal domain. The catalytic domain core of PHDs consists of eight β-strands in a "jelly-roll" or double stranded β helix <scene name='40/400587/Molecular_playground/5'>(DSBH) fold motif</scene> supported by three conserved α-helices and other β-strands and loops that pack along the core. Possession of the DSBH motif is typical of 2-OG-dependent oxygenases.

			The <scene name='40/400587/Active_site/8'>active site</scene>, which is located on a deep cleft between the β-strands comprising the DBSH core, contains the essential Fe(II). It is normally coordinated by the three Fe(II)-binding <scene name='40/400587/Active_site/14'>ligands</scene> formed by the conserved triad sequence, His-X-Asp/Glu-Xn-His.<ref name="review" /><ref>Schofield, C.J., Ratcliffe, P.J. "Signalling Bypoxia by HIF Hydroxylases." Biochemical and Biophysical Research Communications, August 24, 2005, 338, 617-626. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/16139242 16139242]</ref><ref name="structure" /> , 2-OG (which in this scene is replaced by HG) and a water molecule to form an octahedral geometry. Aside from the triad motif residues and those that bind 2-OG, the residues that are predominant inside the active site are nonpolar in nature. This is evidence of the enzyme's need to protect the protein core from oxidation by reactive species that are sometimes generated from iron-related reactions like the Fenton type reaction.<ref name="structure" />

			Upon binding of substrate, the <scene name='40/400587/B2b3_loop/2'>β2β3</scene> loop region of PHD2 goes through a large conformational change (Figure 1). The β2β3 loop moves toward the active site and closes the active site entrance and it is believed to be essential for the catalytic function of PHD2. The β2β3 loop region is mostly conserved in PHDs with minor differences. Mutational studies on the β2β3 loop region showed that this loop is significant for substrate recognition of PHDs<ref>PMID:18063574</ref>.

			[[Image:PHD2.png\|315 px\|thumb\|Fig. 1: Substrate bound (purple) and unbound (green) PHD2. Figure created with PyMol. PDB IDs 2G19, 3HQR.]]

			=== Function ===
			----
			The intrinsic dependence of PHD-catalyzed hydroxylation reactions on molecular oxygen concentration led to the most notable role of PHDs as cellular oxygen sensors. The hydroxylation happens in the oxygen degradation domain at position 4 of the residues Pro-402 (NODD) and Pro-564 (CODD) of [http://en.wikipedia.org/wiki/Hypoxia_(medical) hypoxia]-inducible transcription factor, [http://en.wikipedia.org/wiki/HIF1A (HIF)-α].<ref name="review" />
			Proline hydroxylation in NODD and CODD enable recognition of (HIF)-α by the von Hippel Lindau (pVHL) ubiquitin ligase complex which ultimately results to degradation by the proteasome<ref>PMID:11292862</ref> <ref>PMID:11292861</ref>.

			The requirement of PHDs for the [http://en.wikipedia.org/wiki/Citric_acid_cycle TCA cycle] intermediate, 2-oxoglutarate, also opens the possibility of these enzymes acting as regulators of processes that relate metabolic activity to oxygen levels. Aside from regulation of oxygen homeostasis, other biological functions of the enzyme, which may be hydroxylase-independent or still hydroxylase-dependent but [http://en.wikipedia.org/wiki/HIF1A (HIF)-α]-independent, are being proposed. This is mainly based on the results of various studies: some showed that other factors such as [http://en.wikipedia.org/wiki/Nitric_oxide nitric oxide], [http://en.wikipedia.org/wiki/Reactive_oxygen_species reactive oxygen species] (ROS), and several [http://en.wikipedia.org/wiki/Oncogene oncogenes] control PHD oxygenase activity<ref>Kaelin, W.G. "Proline Hydroxylation and Gene Expression." Annu.Rev.Biochem., February 8, 2005, 74, 115-128. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/15952883 15952883]</ref>; while others described PHD activity on other substrates like [http://en.wikipedia.org/wiki/IKK2 IKK-β]<ref name="review" />. In fact, several functions of the enzyme have been recently identified based on these studies. Listed below are the currently identified functions for PHDs in general<ref name="review" />:
			*tumor suppressor
			*promoter of cell death ([http://en.wikipedia.org/wiki/Apoptosis apoptosis])
			*regulator of cell differentiation

			===3D structures of hypoxia-inducible factor prolyl hydroxylase ===

			[[Hypoxia-inducible factor prolyl hydroxylase]]

			=== References ===
			----
	<references/>		<references/>

Vanessa Chaplin: New page: ==Your Heading Here (maybe something like 'Structure')== This is a default text for you...

2014-11-19T17:34:12Z

New page: ==Your Heading Here (maybe something like 'Structure')== <StructureSection load='1stp' size='340' side='right' caption='Caption for this structure' scene=''> This is a default text for you...

New page

==Your Heading Here (maybe something like 'Structure')==
<StructureSection load='1stp' size='340' side='right' caption='Caption for this structure' scene=''>
This is a default text for your page '''Vanessa Chaplin/sandbox2'''. Click above on '''edit this page''' to modify. Be careful with the < and > signs.
You may include any references to papers as in: the use of JSmol in Proteopedia <ref>DOI 10.1002/ijch.201300024</ref> or to the article describing Jmol <ref>PMID:21638687</ref> to the rescue.

== Function ==

== Disease ==

== Relevance ==

== Structural highlights ==

== Headline text ==

This is a sample scene created with SAT to <scene name="/12/3456/Sample/1">color</scene> by Group, and another to make <scene name="/12/3456/Sample/2">a transparent representation</scene> of the protein. You can make your own scenes on SAT starting from scratch or loading and editing one of these sample scenes.

</StructureSection>
== References ==
<references/>

← Older revision		Revision as of 20:41, 21 November 2014
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	Metazoans adapt to oxygen levels in the environment by making use of intracellular oxygen levels as signals to regulate the [http://en.wikipedia.org/wiki/Transcription_(genetics) transcription] of genes that are essential under normoxic or [http://en.wikipedia.org/wiki/Hypoxia_(medical) hypoxic] conditions. Central to this mechanism is the oxygen-dependent hydroxylation of specific proline and asparagine residues of the transcription factor, hypoxia-inducible factor [http://en.wikipedia.org/wiki/HIF1A (HIF)-α] by the HIF-hydroxylase enzymes, which are studied extensively by the [http://people.chem.umass.edu/knapplab/?q=knappchem/index.html Knapp Lab] here at UMass Amherst.<ref name="review">Fong, G.H., Takeda, K. "Role and Regulation of Prolyl Hydroxylase Domain Proteins." Cell Death and Differentiation, February 15, 2008, 15, 635-641. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/18259202 18259202]</ref>		Metazoans adapt to oxygen levels in the environment by making use of intracellular oxygen levels as signals to regulate the [http://en.wikipedia.org/wiki/Transcription_(genetics) transcription] of genes that are essential under normoxic or [http://en.wikipedia.org/wiki/Hypoxia_(medical) hypoxic] conditions. Central to this mechanism is the oxygen-dependent hydroxylation of specific proline and asparagine residues of the transcription factor, hypoxia-inducible factor [http://en.wikipedia.org/wiki/HIF1A (HIF)-α] by the HIF-hydroxylase enzymes, which are studied extensively by the [http://people.chem.umass.edu/knapplab/?q=knappchem/index.html Knapp Lab] here at UMass Amherst.<ref name="review">Fong, G.H., Takeda, K. "Role and Regulation of Prolyl Hydroxylase Domain Proteins." Cell Death and Differentiation, February 15, 2008, 15, 635-641. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/18259202 18259202]</ref>

	These HIF-hydroxylase enzymes include the Prolyl Hydroxylase Domain isoforms (PHDs) and Factor Inhibition Hypoxia Inducible Factor (FIH). PHDs belong to the same oxygenase superfamily as the [http://en.wikipedia.org/wiki/Collagen collagen] prolyl hydroxylases. Inside the cell, PHD1 is found in the nucleus, while PHD2 is mostly located in the cytoplasm and PHD3 is distributed in both. <ref>PMID:12615973</ref> This distribution contrasts with that of the [http://en.wikipedia.org/wiki/Collagen collagen] prolyl hydroxylases, which reside in the endoplasmic reticulum. In mammals, the PHD dioxygenase subfamily originally included three [http://en.wiktionary.org/wiki/homolog homolog] members: PHD1 (also known as HPH3 and [http://en.wikipedia.org/wiki/EGLN2 EGLN2]), PHD2 (also known as HPH2 and [http://en.wikipedia.org/wiki/EGLN1 EGLN1]), PHD3 (also known as HPH1 and [http://en.wikipedia.org/wiki/EGLN3 EGLN3]). Recently, a newly identified enzyme called P4H-TM (also named PHD4 and EGLN4) was added to this subfamily. Both PHD1 and PHD2 contain more than 400 [http://en.wikipedia.org/wiki/Amino_acid amino acid] residues while PHD3 has less than 250. All isoforms, however, contain the highly conserved hydroxylase domain in the catalytic carboxy-terminal region. <ref name="review" /> PHDs differ in terms of the proline hydroxylation site. PHD2 hydroxylates either the Pro-402 or Pro-564 residue in the oxygen degradation domain of [http://en.wikipedia.org/wiki/HIF1A (HIF)-α] whereas PHD3 can only hydroxylate the Pro-564 position.		These HIF-hydroxylase enzymes include the Prolyl Hydroxylase Domain isoforms (PHDs) and Factor Inhibition Hypoxia Inducible Factor (FIH). PHDs belong to the same oxygenase superfamily as the [http://en.wikipedia.org/wiki/Collagen collagen] prolyl hydroxylases. Inside the cell, PHD1 is found in the nucleus, while PHD2 is mostly located in the cytoplasm and PHD3 is distributed in both. <ref>PMID:12615973</ref> This distribution contrasts with that of the [http://en.wikipedia.org/wiki/Collagen collagen] prolyl hydroxylases, which reside in the endoplasmic reticulum. In mammals, the PHD dioxygenase subfamily originally included three [http://en.wiktionary.org/wiki/homolog homolog] members: PHD1 (also known as HPH3 and [http://en.wikipedia.org/wiki/EGLN2 EGLN2]), PHD2 (also known as HPH2 and [http://en.wikipedia.org/wiki/EGLN1 EGLN1]), PHD3 (also known as HPH1 and [http://en.wikipedia.org/wiki/EGLN3 EGLN3]). Recently, a newly identified enzyme called P4H-TM (also named PHD4 and EGLN4) was added to this subfamily. Both PHD1 and PHD2 contain more than 400 [http://en.wikipedia.org/wiki/Amino_acid amino acid] residues while PHD3 has less than 250. All isoforms, however, contain the highly conserved hydroxylase domain in the catalytic carboxy-terminal region. <ref name="review" /> PHDs differ in terms of the proline hydroxylation site. PHD2 hydroxylates either the Pro-402 or Pro-564 residue in the oxygen degradation domain of [http://en.wikipedia.org/wiki/HIF1A HIF-α] whereas PHD3 can only hydroxylate the Pro-564 position.

	Of the HIF-hydroxylases, '''[http://www.chem.qmul.ac.uk/iubmb/enzyme/EC1/14/11/ (PHD2)]''' is believed to be the main oxygen sensor. PHD2 is a Fe(II)/2-oxoglutarate (OG)-dependent [http://en.wikipedia.org/wiki/Oxygenase dioxygenase] that catalyzes the ''trans''-4-hydroxylation of specific proline residues (as mentioned above) in [http://en.wikipedia.org/wiki/HIF1A (HIF)-α]. In addition to iron, this enzyme also requires [http://en.wikipedia.org/wiki/Vitamin_C ascorbate] as a cofactor.<ref name="structure">Mcdonough, M.A., Li, V., Flashman, E., et al. "Cellular oxygen sensing: Crystal structure of hypoxia-inducible factor prolyl hydroxylase (PHD2)." PNAS, June 27, 2006, 103 (26), 9814-9819. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/16782814 16782814]</ref> Our goal in the Knapp Lab is to understand and define the mechanism of action for this enzyme and evaluate how oxygen affects enzyme activity to ~~gaining further~~ knowledge of oxygen sensing.		Of the HIF-hydroxylases, '''[http://www.chem.qmul.ac.uk/iubmb/enzyme/EC1/14/11/ PHD2]''' is believed to be the main oxygen sensor. PHD2 is a Fe(II)/2-oxoglutarate (OG)-dependent [http://en.wikipedia.org/wiki/Oxygenase dioxygenase] that catalyzes the ''trans''-4-hydroxylation of specific proline residues (as mentioned above) in [http://en.wikipedia.org/wiki/HIF1A HIF-α]. In addition to iron, this enzyme also requires [http://en.wikipedia.org/wiki/Vitamin_C ascorbate] as a cofactor.<ref name="structure">Mcdonough, M.A., Li, V., Flashman, E., et al. "Cellular oxygen sensing: Crystal structure of hypoxia-inducible factor prolyl hydroxylase (PHD2)." PNAS, June 27, 2006, 103 (26), 9814-9819. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/16782814 16782814]</ref> Our goal in the Knapp Lab is to understand and define the mechanism of action for this enzyme and evaluate how oxygen affects enzyme activity to improve knowledge of oxygen sensing.

	<structure load='2g19' size='335' frame='true' align='right'		<structure load='2g19' size='335' frame='true' align='right'
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	<scene name='Molecular_Playground/Prolyl_Hydroxylase_Domain_(PHD)_Enzyme/Molecular_playground/4'>Molecular Playground</scene>		<scene name='Molecular_Playground/Prolyl_Hydroxylase_Domain_(PHD)_Enzyme/Molecular_playground/4'>Molecular Playground</scene>

	Molecular Playground banner: Prolyl Hydroxylase Domain (~~PHD~~) enzyme, a cellular oxygen sensor, has a major regulatory role in oxygen homeostasis.		Molecular Playground banner: Prolyl Hydroxylase Domain 2 (PHD2) enzyme, a cellular oxygen sensor, has a major regulatory role in oxygen homeostasis.

	=== Structure ===		=== Structure ===
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	PHDs have two structural domains: the more variable N-terminal domain and the conserved catalytic C-terminal domain. The catalytic domain core of PHDs consists of eight β-strands in a "jelly-roll" or double stranded β helix <scene name='40/400587/Molecular_playground/5'>(DSBH) fold motif</scene> supported by three conserved α-helices and other β-strands and loops that pack along the core. Possession of the DSBH motif is typical of 2-OG-dependent oxygenases.		PHDs have two structural domains: the more variable N-terminal domain and the conserved catalytic C-terminal domain. The catalytic domain core of PHDs consists of eight β-strands in a "jelly-roll" or double stranded β helix <scene name='40/400587/Molecular_playground/5'>(DSBH) fold motif</scene> supported by three conserved α-helices and other β-strands and loops that pack along the core. Possession of the DSBH motif is typical of 2-OG-dependent oxygenases.

	The <scene name='40/400587/Active_site/8'>active site</scene>, which is located on a deep cleft between the β-strands comprising the DBSH core, contains the essential Fe(II). It is normally coordinated by the three Fe(II)-binding <scene name='40/400587/Active_site/14'>ligands</scene> formed by the conserved triad sequence, His-X-Asp/Glu-Xn-His.<ref name="review" /><ref>Schofield, C.J., Ratcliffe, P.J. "Signalling Bypoxia by HIF Hydroxylases." Biochemical and Biophysical Research Communications, August 24, 2005, 338, 617-626. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/16139242 16139242]</ref><ref name="structure" /> , 2-OG (which in this scene is replaced by HG) and a water molecule to form an octahedral geometry. Aside from the triad motif residues and those that bind 2-OG, the residues that are predominant inside the active site are nonpolar in nature. This is evidence of the enzyme's need to protect the protein core from oxidation by reactive species that are sometimes generated from iron-related reactions ~~like the~~ Fenton ~~type reaction~~.<ref name="structure" />		The <scene name='40/400587/Active_site/8'>active site</scene>, which is located on a deep cleft between the β-strands comprising the DBSH core, contains the essential Fe(II). It is normally coordinated by the three Fe(II)-binding <scene name='40/400587/Active_site/14'>ligands</scene> formed by the conserved triad sequence, His-X-Asp/Glu-Xn-His.<ref name="review" /><ref>Schofield, C.J., Ratcliffe, P.J. "Signalling Bypoxia by HIF Hydroxylases." Biochemical and Biophysical Research Communications, August 24, 2005, 338, 617-626. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/16139242 16139242]</ref><ref name="structure" /> , 2-OG (which in this scene is replaced by HG) and a water molecule to form an octahedral geometry. Aside from the triad motif residues and those that bind 2-OG, the residues that are predominant inside the active site are nonpolar in nature. This is evidence of the enzyme's need to protect the protein core from oxidation by reactive species that are sometimes generated from iron-related reactions such as Fenton chemistry.<ref name="structure" />

	Upon binding of substrate, the <scene name='40/400587/B2b3_loop/2'>β2β3</scene> loop region of PHD2 ~~goes through~~ a large conformational change (Figure 1). The β2β3 loop moves toward the active site and closes the active site entrance ~~and it~~ is believed to be essential for the catalytic function of PHD2. The β2β3 loop region is mostly conserved in PHDs with minor differences. Mutational studies on the β2β3 loop region showed that this loop is significant for substrate recognition of PHDs<ref>PMID:18063574</ref>.		Upon binding of substrate, the <scene name='40/400587/B2b3_loop/2'>β2β3</scene> loop region of PHD2 undergoes a large conformational change (Figure 1). The β2β3 loop moves toward the active site and closes over the active site entrance. This loop movement is believed to be essential for the catalytic function of PHD2. The β2β3 loop region is mostly conserved in PHDs with minor differences. Mutational studies on the β2β3 loop region showed that this loop is significant for substrate recognition of PHDs<ref>PMID:18063574</ref>.

	[[Image:PHD2.png\|315 px\|thumb\|Fig. 1: Substrate bound (purple) and unbound (green) PHD2. Figure created with PyMol. PDB IDs 2G19, 3HQR.]]		[[Image:PHD2.png\|315 px\|thumb\|Fig. 1: Substrate bound (purple) and unbound (green) PHD2. Figure created with PyMol. PDB IDs 2G19, 3HQR.]]
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	=== Function ===		=== Function ===
	----		----
	The intrinsic dependence of PHD-catalyzed hydroxylation reactions on molecular oxygen concentration led to the most notable role of PHDs as cellular oxygen sensors. ~~The hydroxylation happens in~~ the oxygen degradation domain at position 4 of the residues Pro-402 (NODD) and Pro-564 (CODD) of [http://en.wikipedia.org/wiki/Hypoxia_(medical) hypoxia]-inducible transcription factor, [http://en.wikipedia.org/wiki/HIF1A (HIF)-α].<ref name="review" />		The intrinsic dependence of PHD-catalyzed hydroxylation reactions on molecular oxygen concentration led to the most notable role of PHDs as cellular oxygen sensors. Hydroxylation occurs within the oxygen degradation domain at position 4 of the residues Pro-402 (NODD) and Pro-564 (CODD) of [http://en.wikipedia.org/wiki/Hypoxia_(medical) hypoxia]-inducible transcription factor, [http://en.wikipedia.org/wiki/HIF1A HIF-α].<ref name="review" />
	Proline hydroxylation in NODD and CODD enable recognition of (HIF)-α by the von Hippel Lindau (pVHL) ubiquitin ligase complex which ultimately results to degradation by the proteasome<ref>PMID:11292862</ref> <ref>PMID:11292861</ref>.		Proline hydroxylation in NODD and CODD enable recognition of HIF-α by the von Hippel Lindau (pVHL) ubiquitin ligase complex which ultimately results in degradation of HIF-α by the proteasome<ref>PMID:11292862</ref> <ref>PMID:11292861</ref>.

	The requirement of PHDs for the [http://en.wikipedia.org/wiki/Citric_acid_cycle TCA cycle] intermediate, 2-oxoglutarate, also opens the possibility of these enzymes acting as regulators of processes that relate metabolic activity to oxygen levels. Aside from regulation of oxygen homeostasis, other biological functions of the enzyme, which may be hydroxylase-independent or still hydroxylase-dependent but [http://en.wikipedia.org/wiki/HIF1A (HIF)-α]-independent, are being proposed. This is mainly based on the results of various studies: some showed that other factors such as [http://en.wikipedia.org/wiki/Nitric_oxide nitric oxide], [http://en.wikipedia.org/wiki/Reactive_oxygen_species reactive oxygen species] (ROS), and several [http://en.wikipedia.org/wiki/Oncogene oncogenes] control PHD oxygenase activity<ref>Kaelin, W.G. "Proline Hydroxylation and Gene Expression." Annu.Rev.Biochem., February 8, 2005, 74, 115-128. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/15952883 15952883]</ref>; while others described PHD activity on other substrates like [http://en.wikipedia.org/wiki/IKK2 IKK-β]<ref name="review" />. In fact, several functions of the enzyme have been recently identified based on these studies. Listed below are the currently identified functions for PHDs in general<ref name="review" />:		The requirement of PHDs for the [http://en.wikipedia.org/wiki/Citric_acid_cycle TCA cycle] intermediate, 2-oxoglutarate, also opens the possibility of these enzymes acting as regulators of processes that relate metabolic activity to oxygen levels. Aside from regulation of oxygen homeostasis, other biological functions of the enzyme, which may be hydroxylase-independent or still hydroxylase-dependent but [http://en.wikipedia.org/wiki/HIF1A HIF-α]-independent, are being proposed. This is mainly based on the results of various studies: some showed that other factors such as [http://en.wikipedia.org/wiki/Nitric_oxide nitric oxide], [http://en.wikipedia.org/wiki/Reactive_oxygen_species reactive oxygen species] (ROS), and several [http://en.wikipedia.org/wiki/Oncogene oncogenes] control PHD oxygenase activity<ref>Kaelin, W.G. "Proline Hydroxylation and Gene Expression." Annu.Rev.Biochem., February 8, 2005, 74, 115-128. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/15952883 15952883]</ref>; while others described PHD activity on other substrates like [http://en.wikipedia.org/wiki/IKK2 IKK-β]<ref name="review" />. In fact, several functions of the enzyme have been recently identified based on these studies. Listed below are the currently identified functions for PHDs in general<ref name="review" />:
	*tumor suppressor		*tumor suppressor
	*promoter of cell death ([http://en.wikipedia.org/wiki/Apoptosis apoptosis])		*promoter of cell death ([http://en.wikipedia.org/wiki/Apoptosis apoptosis])