Proteopedia - User contributions [en]

Phosphoglycerate Kinase

2010-03-31T18:08:00Z

Shane Harmon: /* Phosphoglycerate Kinase(PGK) */

=Phosphoglycerate Kinase(PGK)=

== PGK in the Glycolysis Cycle ==

Phosphoglycerate kinase is a crucial enzyme in the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphoglycerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/2'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/4'>ATP</scene>. This method for ATP production is known as substrate-level phosphorylation because it produces energy storing ATP molecules without the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously.

{{STRUCTURE_3cin | PDB=1PHP | SCENE= }}

== Structure ==

The overall structure of Phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of approximately 400 amino acids, with a molecular weight of about 45kD <ref> Auerbach, Gunter et al. 1997. Closed Structure of phosphoglycerate kinase from Thermotoga maritima reveals the catalytic mechanism and determinants of thermal stability. Structure. 5:1475-1483.</ref>. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations: <scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP classification of PGK is alpha and beta, indicating that its <scene name='Shane_Harmon_Sandbox/Scop_classifcation/1'>secondary structure</scene>is composed of roughly equal numbers alpha and beta sheets.

== Reaction Mechanism ==

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. The N-terminal domain has a basic region where the 1,3-Biphosphoglycerate and 3-phosphoglycerate bind while the C-terminal domain has the binding sites for the nucleotide substrates, ADP and ATP. Upon binding of both substrate molecules at the active sites, the protein’s conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. This hydrophobic chamber is necessary to prevent ATP hydrolysis <ref> Auerbach, Gunter et al. 1997. Closed Structure of phosphoglycerate kinase from Thermotoga maritima reveals the catalytic mechanism and determinants of thermal stability. Structure. 5:1475-1483.</ref> The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Arg_and_asp/2'>Arg 62 and Asp 200</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> Recent research indicates that the mechanism for closure of the two domains is a series of hydrogen bond interactions that occur upon binding of the substrates on both domains <ref>Vas, M, Varga, A et al. 2010. Insight into the Mechanism of of Domain Movements and their Role in Enzyme Function: Example of 3-Phosphoglycerate kinase. Current Protein and Peptide Science. Jan 21, 2010. (Epub ahead of publication).</ref>

The mechanism of catalysis has not been fully established because the PGK/1-3biphophoglycerate complex is highly unstable; however, it is thought that the mechanism is similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes the C1 phosphate group from 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref>

The general mechanism is a single displacement Sn2 reaction in which the ADP-B-phosphate oxygen atom initiates nucleophilic attack on the 1-phosphate group of 1-3biphosphoglycerate <ref> Auerbach, Gunter et al. 1997. Closed Structure of phosphoglycerate kinase from Thermotoga maritima reveals the catalytic mechanism and determinants of thermal stability. Structure. 5:1475-1483.</ref>. Thus, the phosphoryl group is transferred directly via a charged transition state. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form <scene name='Shane_Harmon_Sandbox/Atp/5'>hydrogen bonds</scene> with the enzyme. The 3 hydrogen bonds of ATP are favored over the 2 hydrogen bonds of ADP.

[[Image:PGKmechanism2.jpg]]

Two specific residues known to be necessary for catalysis are <scene name='Shane_Harmon_Sandbox/197_and_38/2'>Lys 197 and Arg 36</scene>. Lys 197 secures 1,3-biphosphoblycerate in the closed conformation, and it has been proposed that the transition state intermediary is stabilized by the highly conserved Lys 197 as it transfers the phosphate group. Additionally, it has been shown that Arg 38 is also necessary for catalytic function. Arg 36 has been shown to stabilize a water molecule in the closed conformation and may form a hydrogen bond with the ATP product <ref> Auerbach, Gunter et al. 1997. Closed Structure of phosphoglycerate kinase from Thermotoga maritima reveals the catalytic mechanism and determinants of thermal stability. Structure. 5:1475-1483.</ref>.

== Kinetics ==

Given that Phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-Hofstee plots curve upward. One possible explanation for the non-linearity, negative cooperativity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 </ref>

== Regulation ==

The role of PGK in glycolysis is very important for the production of ATP through substrate level phosphorylation. Regulation of this protein is thereby controlled by ATP or energy level of the cell. Recent research in frogs which can withstand freezing temperatures indicates that PGK is up regulated by the cold and more specifically by low levels of oxygen <ref> Shaobo, Wu et al. 2009. PGK1 expression responds to freezing, anoxia, and dehydration stresses in freeze tolerant wood frog, Rana sylvatica. Journal of Experimental Zoology. 311, 57-67 </ref> This makes logical sense as when oxygen is low, oxidative phosphorylation cannot occur. Thus without oxygen and oxidative phosphorylation, ATP levels begin to drop. In response to decreased ATP, PGK is up regulated to increase the amount of ATP produced by substrate level phosphorylation. It could therefore also be expected that ADP might act to inhibit or down regulated PGK expression.

== Other Functions ==

Recent study of PGK has revolved around its function in tumor formation and growth. It has been shown that in addition to catalyzing its normal reaction of 1,3-Biphosphoglycerate and ADP to ATP and 3-Phosphoglycerate, PGK can also function to cleave disulfide bonds. Specifically, the review of sulfide bond cleavage indicates PGK has been shown to cleave disulfide bonds in the protein zymogen plasmin to produce the active form of the protein. The active form of plasmin is responsible for angiogenesis or blood vessel formation in tumors <ref> Hogg, PJ. 2002. Biological Regulation through protein disulfide bond cleavage. Redox Report. 7(2), 71-77. </ref> Without the formation of blood vessels in tumors, nutrients are limited and tumor growth is therfore limited. Once blood vessels are established growth can rapidly increase. The fact that tumor cells secrete PGK to allow blood vessel formation through the activation of the zymogen plasmin has important implications for understanding its regulation. If the regulation of PGK in tumor cells can be understood, it might be possible to inhibit the overproduction and secretion of PGK to limit angiogenesis in tumors.

<references/>

Phosphoglycerate Kinase

2010-03-31T18:02:04Z

Shane Harmon: /* Phosphoglycerate Kinase(PGK) */

=Phosphoglycerate Kinase(PGK)=

== PGK in the Glycolysis Cycle ==

Phosphoglycerate kinase is a crucial enzyme in the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphoglycerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/2'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/4'>ATP</scene>. This method for ATP production is known as substrate-level phosphorylation because it produces energy storing ATP molecules without the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously.

{{STRUCTURE_3cin | PDB=1PHP | SCENE= }}

== Structure ==

The overall structure of Phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of approximately 400 amino acids, with a molecular weight of about 45kD <ref> Auerbach, Gunter et al. 1997. Closed Structure of phosphoglycerate kinase from Thermotoga maritima reveals the catalytic mechanism and determinants of thermal stability. Structure. 5:1475-1483.</ref>. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations: <scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP classification of PGK is alpha and beta, indicating that its <scene name='Shane_Harmon_Sandbox/Scop_classifcation/1'>secondary structure</scene>is composed of roughly equal numbers alpha and beta sheets.

== Reaction Mechanism ==

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. The N-terminal domain has a basic region where the 1,3-Biphosphoglycerate and 3-phosphoglycerate bind while the C-terminal domain has the binding sites for the nucleotide substrates, ADP and ATP. Upon binding of both substrate molecules at the active sites, the protein’s conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. This hydrophobic chamber is necessary to prevent ATP hydrolysis <ref> Auerbach, Gunter et al. 1997. Closed Structure of phosphoglycerate kinase from Thermotoga maritima reveals the catalytic mechanism and determinants of thermal stability. Structure. 5:1475-1483.</ref> The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Arg_and_asp/2'>Arg 62 and Asp 200</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> Recent research indicates that the mechanism for closure of the two domains is a series of hydrogen bond interactions that occur upon binding of the substrates on both domains <ref>Vas, M, Varga, A et al. 2010. Insight into the Mechanism of of Domain Movements and their Role in Enzyme Function: Example of 3-Phosphoglycerate kinase. Current Protein and Peptide Science. Jan 21, 2010. (Epub ahead of publication).</ref>

The mechanism of catalysis has not been fully established because the PGK/1-3biphophoglycerate complex is highly unstable; however, it is thought that the mechanism is similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes the C1 phosphate group from 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref>

The general mechanism is a single displacement Sn2 reaction in which the ADP-B-phosphate oxygen atom initiates nucleophilic attack on the 1-phosphate group of 1-3biphosphoglycerate <ref> Auerbach, Gunter et al. 1997. Closed Structure of phosphoglycerate kinase from Thermotoga maritima reveals the catalytic mechanism and determinants of thermal stability. Structure. 5:1475-1483.</ref>. Thus, the phosphoryl group is transferred directly via a charged transition state. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form <scene name='Shane_Harmon_Sandbox/Atp/5'>hydrogen bonds</scene> with the enzyme. The 3 hydrogen bonds of ATP are favored over the 2 hydrogen bonds of ADP.

[[Image:PGKmechanism2.jpg]]

Two specific residues known to be necessary for catalysis are <scene name='Shane_Harmon_Sandbox/197_and_38/2'>Lys 197 and Arg 36</scene>. Lys 197 secures 1,3-biphosphoblycerate in the closed conformation, and it has been proposed that the transition state intermediary is stabilized by the highly conserved Lys 197 as it transfers the phosphate group. Additionally, it has been shown that Arg 38 is also necessary for catalytic function. Arg 36 has been shown to stabilize a water molecule in the closed conformation and may form a hydrogen bond with the ATP product <ref> Auerbach, Gunter et al. 1997. Closed Structure of phosphoglycerate kinase from Thermotoga maritima reveals the catalytic mechanism and determinants of thermal stability. Structure. 5:1475-1483.</ref>.

== Kinetics ==

Given that Phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-Hofstee plots curve upward. One possible explanation for the non-linearity, negative cooperativity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 </ref>

== Regulation ==

The role of PGK in glycolysis is very important for the production of ATP through substrate level phosphorylation. Regulation of this protein is thereby controlled by ATP or energy level of the cell. Recent research in frogs which can withstand freezing temperatures indicates that PGK is up regulated by the cold and more specifically by low levels of oxygen <ref> Shaobo, Wu et al. 2009. PGK1 expression responds to freezing, anoxia, and dehydration stresses in freeze tolerant wood frog, Rana sylvatica. Journal of Experimental Zoology. 311, 57-67 </ref> This makes logical sense as when oxygen is low, oxidative phosphorylation cannot occur. Thus without oxygen and oxidative phosphorylation, ATP levels begin to drop. In response to decreased ATP, PGK is up regulated to increase the amount of ATP produced by substrate level phosphorylation. It could therefore also be expected that ADP might act to inhibit or down regulated PGK expression.

Recent study of PGK has revolved around its function in tumor formation and growth. It has been shown that in addition to catalyzing its normal reaction of 1,3-Biphosphoglycerate and ADP to ATP and 3-Phosphoglycerate, PGK can also function to cleave disulfide bonds. Specifically, the review of sulfide bond cleavage indicates PGK has been shown to cleave disulfide bonds in the protein zymogen plasmin to produce the active form of the protein. The active form of plasmin is responsible for angiogenesis or blood vessel formation in tumors <ref> Hogg, PJ. 2002. Biological Regulation through protein disulfide bond cleavage. Redox Report. 7(2), 71-77. </ref> Without the formation of blood vessels to tumors nutrients are limited and tumor growth is limited. Once blood vessels are established growth can rapidly increase. The fact that tumor cells secrete PGK to allow blood vessel formation through the activation of the zymogen plasmin has important implications for understanding its regulation. If the regulation of PGK in tumor cells can be understood, it might be possible to inhibit the overproduction and secretion of PGK to limit angiogenesis in tumors.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-31T18:00:24Z

Shane Harmon: /* Phosphoglycerate Kinase(PGK) */

=Phosphoglycerate Kinase(PGK)=

== PGK in the Glycolysis Cycle ==

Phosphoglycerate kinase is a crucial enzyme in the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphoglycerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/2'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/4'>ATP</scene>. This method for ATP production is known as substrate-level phosphorylation because it produces energy storing ATP molecules without the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously.

{{STRUCTURE_3cin | PDB=1PHP | SCENE= }}

== Structure ==

The overall structure of Phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of approximately 400 amino acids, with a molecular weight of about 45kD <ref> Auerbach, Gunter et al. 1997. Closed Structure of phosphoglycerate kinase from Thermotoga maritima reveals the catalytic mechanism and determinants of thermal stability. Structure. 5:1475-1483.</ref>. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations: <scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP classification of PGK is alpha and beta, indicating that its <scene name='Shane_Harmon_Sandbox/Scop_classifcation/1'>secondary structure</scene>is composed of roughly equal numbers alpha and beta sheets.

== Reaction Mechanism ==

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. The N-terminal domain has a basic region where the 1,3-Biphosphoglycerate and 3-phosphoglycerate bind while the C-terminal domain has the binding sites for the nucleotide substrates, ADP and ATP. Upon binding of both substrate molecules at the active sites, the protein’s conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. This hydrophobic chamber is necessary to prevent ATP hydrolysis <ref> Auerbach, Gunter et al. 1997. Closed Structure of phosphoglycerate kinase from Thermotoga maritima reveals the catalytic mechanism and determinants of thermal stability. Structure. 5:1475-1483.</ref> The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Arg_and_asp/2'>Arg 62 and Asp 200</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> Recent research indicates that the mechanism for closure of the two domains is a series of hydrogen bond interactions that occur upon binding of the substrates on both domains <ref>Vas, M, Varga, A et al. 2010. Insight into the Mechanism of of Domain Movements and their Role in Enzyme Function: Example of 3-Phosphoglycerate kinase. Current Protein and Peptide Science. Jan 21, 2010. (Epub ahead of publication).</ref>

The mechanism of catalysis has not been fully established because the PGK/1-3biphophoglycerate complex is highly unstable; however, it is thought that the mechanism is similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes the C1 phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref>

The general mechanism is a single displacement Sn2 reaction in which the ADP-B-phosphate oxygen atom initiates nucleophilic attack on the 1-phosphate group of 1-3biphosphoglycerate <ref> Auerbach, Gunter et al. 1997. Closed Structure of phosphoglycerate kinase from Thermotoga maritima reveals the catalytic mechanism and determinants of thermal stability. Structure. 5:1475-1483.</ref>. Thus, the phosphoryl group is transferred directly via a charged transition state. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form <scene name='Shane_Harmon_Sandbox/Atp/5'>hydrogen bonds</scene> with the enzyme. The 3 hydrogen bonds of ATP are favored over the 2 hydrogen bonds of ADP.

[[Image:PGKmechanism2.jpg]]

Two specific residues known to be necessary for catalysis are <scene name='Shane_Harmon_Sandbox/197_and_38/2'>Lys 197 and Arg 36</scene>. Lys 197 secures 1,3-biphosphoblycerate in the closed conformation, and it has been proposed that the transition state intermediary is stabilized by the highly conserved Lys 197 as it transfers the phosphate group. Additionally, it has been shown that Arg 38 is also necessary for catalytic function. Arg 36 has been shown to stabilize a water molecule in the closed conformation and may form a hydrogen bond with the ATP product <ref> Auerbach, Gunter et al. 1997. Closed Structure of phosphoglycerate kinase from Thermotoga maritima reveals the catalytic mechanism and determinants of thermal stability. Structure. 5:1475-1483.</ref>.

== Kinetics ==

Given that Phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-Hofstee plots curve upward. One possible explanation for the non-linearity, negative cooperativity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 </ref>

== Regulation ==

The role of PGK in glycolysis is very important for the production of ATP through substrate level phosphorylation. Regulation of this protein is thereby controlled by ATP or energy level of the cell. Recent research in frogs which can withstand freezing temperatures indicates that PGK is up regulated by the cold and more specifically by low levels of oxygen <ref> Shaobo, Wu et al. 2009. PGK1 expression responds to freezing, anoxia, and dehydration stresses in freeze tolerant wood frog, Rana sylvatica. Journal of Experimental Zoology. 311, 57-67 </ref> This makes logical sense as when oxygen is low, oxidative phosphorylation cannot occur. Thus without oxygen and oxidative phosphorylation, ATP levels begin to drop. In response to decreased ATP, PGK is up regulated to increase the amount of ATP produced by substrate level phosphorylation. It could therefore also be expected that ADP might act to inhibit or down regulated PGK expression.

Recent study of PGK has revolved around its function in tumor formation and growth. It has been shown that in addition to catalyzing its normal reaction of 1,3-Biphosphoglycerate and ADP to ATP and 3-Phosphoglycerate, PGK can also function to cleave disulfide bonds. Specifically, the review of sulfide bond cleavage indicates PGK has been shown to cleave disulfide bonds in the protein zymogen plasmin to produce the active form of the protein. The active form of plasmin is responsible for angiogenesis or blood vessel formation in tumors <ref> Hogg, PJ. 2002. Biological Regulation through protein disulfide bond cleavage. Redox Report. 7(2), 71-77. </ref> Without the formation of blood vessels to tumors nutrients are limited and tumor growth is limited. Once blood vessels are established growth can rapidly increase. The fact that tumor cells secrete PGK to allow blood vessel formation through the activation of the zymogen plasmin has important implications for understanding its regulation. If the regulation of PGK in tumor cells can be understood, it might be possible to inhibit the overproduction and secretion of PGK to limit angiogenesis in tumors.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-31T17:59:33Z

Shane Harmon: /* Phosphoglycerate Kinase(PGK) */

=Phosphoglycerate Kinase(PGK)=

== PGK in the Glycolysis Cycle ==

Phosphoglycerate kinase is a crucial enzyme in the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphoglycerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/2'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/4'>ATP</scene>. This method for ATP production is known as substrate-level phosphorylation because it produces energy storing ATP molecules without the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously.

{{STRUCTURE_3cin | PDB=1PHP | SCENE= }}

== Structure ==

The overall structure of Phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of approximately 400 amino acids, with a molecular weight of about 45kD <ref> Auerbach, Gunter et al. 1997. Closed Structure of phosphoglycerate kinase from Thermotoga maritima reveals the catalytic mechanism and determinants of thermal stability. Structure. 5:1475-1483.</ref>. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations: <scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP classification of PGK is alpha and beta, indicating that its <scene name='Shane_Harmon_Sandbox/Scop_classifcation/1'>secondary structure</scene>is composed of roughly equal numbers alpha and beta sheets.

== Reaction Mechanism ==

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. The N-terminal domain has a basic region where the 1,3-Biphosphoglycerate and 3-phosphoglycerate bind while the C-terminal domain has the binding sites for the nucleotide substrates, ADP and ATP. Upon binding of both substrate molecules at the active sites, the protein’s conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. This hydrophobic chamber is necessary to prevent ATP hydrolysis <ref> Auerbach, Gunter et al. 1997. Closed Structure of phosphoglycerate kinase from Thermotoga maritima reveals the catalytic mechanism and determinants of thermal stability. Structure. 5:1475-1483.</ref> The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Arg_and_asp/2'>Arg 62 and Asp 200</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> Recent research indicates that the mechanism for closure of the two domains is a series of hydrogen bond interactions that occur upon binding of the substrates on both domains <ref>Vas, M, Varga, A et al. 2010. Insight into the Mechanism of of Domain Movements and their Role in Enzyme Function: Example of 3-Phosphoglycerate kinase. Current Protein and Peptide Science. Jan 21, 2010. (Epub ahead of publication).</ref>

The mechanism of catalysis has not been fully established because the PGK/1-3biphophoglycerate complex is highly unstable; however, it is thought that the mechanism is similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes the C1 phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref>

The general mechanism is a single displacement Sn2 reaction in which the ADP-B-phosphate oxygen atom initiates nucleophilic attack on the 1-phosphate group of 1-3biphosphoglycerate. Thus, the phosphoryl group is transferred directly via a charged transition state. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form <scene name='Shane_Harmon_Sandbox/Atp/5'>hydrogen bonds</scene> with the enzyme. The 3 hydrogen bonds of ATP are favored over the 2 hydrogen bonds of ADP.

[[Image:PGKmechanism2.jpg]]

Two specific residues known to be necessary for catalysis are <scene name='Shane_Harmon_Sandbox/197_and_38/2'>Lys 197 and Arg 36</scene>. Lys 197 secures 1,3-biphosphoblycerate in the closed conformation, and it has been proposed that the transition state intermediary is stabilized by the highly conserved Lys 197 as it transfers the phosphate group. Additionally, it has been shown that Arg 38 is also necessary for catalytic function. Arg 36 has been shown to stabilize a water molecule in the closed conformation and may form a hydrogen bond with the ATP product.

== Kinetics ==

Given that Phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-Hofstee plots curve upward. One possible explanation for the non-linearity, negative cooperativity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 </ref>

== Regulation ==

The role of PGK in glycolysis is very important for the production of ATP through substrate level phosphorylation. Regulation of this protein is thereby controlled by ATP or energy level of the cell. Recent research in frogs which can withstand freezing temperatures indicates that PGK is up regulated by the cold and more specifically by low levels of oxygen <ref> Shaobo, Wu et al. 2009. PGK1 expression responds to freezing, anoxia, and dehydration stresses in freeze tolerant wood frog, Rana sylvatica. Journal of Experimental Zoology. 311, 57-67 </ref> This makes logical sense as when oxygen is low, oxidative phosphorylation cannot occur. Thus without oxygen and oxidative phosphorylation, ATP levels begin to drop. In response to decreased ATP, PGK is up regulated to increase the amount of ATP produced by substrate level phosphorylation. It could therefore also be expected that ADP might act to inhibit or down regulated PGK expression.

Recent study of PGK has revolved around its function in tumor formation and growth. It has been shown that in addition to catalyzing its normal reaction of 1,3-Biphosphoglycerate and ADP to ATP and 3-Phosphoglycerate, PGK can also function to cleave disulfide bonds. Specifically, the review of sulfide bond cleavage indicates PGK has been shown to cleave disulfide bonds in the protein zymogen plasmin to produce the active form of the protein. The active form of plasmin is responsible for angiogenesis or blood vessel formation in tumors <ref> Hogg, PJ. 2002. Biological Regulation through protein disulfide bond cleavage. Redox Report. 7(2), 71-77. </ref> Without the formation of blood vessels to tumors nutrients are limited and tumor growth is limited. Once blood vessels are established growth can rapidly increase. The fact that tumor cells secrete PGK to allow blood vessel formation through the activation of the zymogen plasmin has important implications for understanding its regulation. If the regulation of PGK in tumor cells can be understood, it might be possible to inhibit the overproduction and secretion of PGK to limit angiogenesis in tumors.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-31T17:53:55Z

Shane Harmon: /* Phosphoglycerate Kinase(PGK) */

=Phosphoglycerate Kinase(PGK)=

== PGK in the Glycolysis Cycle ==

Phosphoglycerate kinase is a crucial enzyme in the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphoglycerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/2'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/4'>ATP</scene>. This method for ATP production is known as substrate-level phosphorylation because it produces energy storing ATP molecules without the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously.

{{STRUCTURE_3cin | PDB=1PHP | SCENE= }}

== Structure ==

The overall structure of Phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of approximately 400 amino acids, with a molecular weight of about 45kD. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations: <scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP classification of PGK is alpha and beta, indicating that its <scene name='Shane_Harmon_Sandbox/Scop_classifcation/1'>secondary structure</scene>is composed of roughly equal numbers alpha and beta sheets.

== Reaction Mechanism ==

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. The N-terminal domain has a basic region where the 1,3-Biphosphoglycerate and 3-phosphoglycerate bind while the C-terminal domain has the binding sites for the nucleotide substrates, ADP and ATP. Upon binding of both substrate molecules at the active sites, the protein’s conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. This hydrophobic chamber is necessary to prevent ATP hydrolysis <ref> Auerbach, Gunter et al. 1997. Closed Structure of phosphoglycerate kinase from Thermotoga maritima reveals the catalytic mechanism and determinants of thermal stability. Structure. 5:1475-1483.</ref> The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Arg_and_asp/2'>Arg 62 and Asp 200</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> Recent research indicates that the mechanism for closure of the two domains is a series of hydrogen bond interactions that occur upon binding of the substrates on both domains <ref>Vas, M, Varga, A et al. 2010. Insight into the Mechanism of of Domain Movements and their Role in Enzyme Function: Example of 3-Phosphoglycerate kinase. Current Protein and Peptide Science. Jan 21, 2010. (Epub ahead of publication).</ref>

The mechanism of catalysis has not been fully established because the PGK/1-3biphophoglycerate complex is highly unstable; however, it is thought that the mechanism is similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes the C1 phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref>

The general mechanism is a single displacement Sn2 reaction in which the ADP-B-phosphate oxygen atom initiates nucleophilic attack on the 1-phosphate group of 1-3biphosphoglycerate. Thus, the phosphoryl group is transferred directly via a charged transition state. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form <scene name='Shane_Harmon_Sandbox/Atp/5'>hydrogen bonds</scene> with the enzyme. The 3 hydrogen bonds of ATP are favored over the 2 hydrogen bonds of ADP.

[[Image:PGKmechanism2.jpg]]

Two specific residues known to be necessary for catalysis are <scene name='Shane_Harmon_Sandbox/197_and_38/2'>Lys 197 and Arg 36</scene>. Lys 197 secures 1,3-biphosphoblycerate in the closed conformation, and it has been proposed that the transition state intermediary is stabilized by the highly conserved Lys 197 as it transfers the phosphate group. Additionally, it has been shown that Arg 38 is also necessary for catalytic function. Arg 36 has been shown to stabilize a water molecule in the closed conformation and may form a hydrogen bond with the ATP product.

== Kinetics ==

Given that Phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-Hofstee plots curve upward. One possible explanation for the non-linearity, negative cooperativity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 </ref>

== Regulation ==

The role of PGK in glycolysis is very important for the production of ATP through substrate level phosphorylation. Regulation of this protein is thereby controlled by ATP or energy level of the cell. Recent research in frogs which can withstand freezing temperatures indicates that PGK is up regulated by the cold and more specifically by low levels of oxygen <ref> Shaobo, Wu et al. 2009. PGK1 expression responds to freezing, anoxia, and dehydration stresses in freeze tolerant wood frog, Rana sylvatica. Journal of Experimental Zoology. 311, 57-67 </ref> This makes logical sense as when oxygen is low, oxidative phosphorylation cannot occur. Thus without oxygen and oxidative phosphorylation, ATP levels begin to drop. In response to decreased ATP, PGK is up regulated to increase the amount of ATP produced by substrate level phosphorylation. It could therefore also be expected that ADP might act to inhibit or down regulated PGK expression.

Recent study of PGK has revolved around its function in tumor formation and growth. It has been shown that in addition to catalyzing its normal reaction of 1,3-Biphosphoglycerate and ADP to ATP and 3-Phosphoglycerate, PGK can also function to cleave disulfide bonds. Specifically, the review of sulfide bond cleavage indicates PGK has been shown to cleave disulfide bonds in the protein zymogen plasmin to produce the active form of the protein. The active form of plasmin is responsible for angiogenesis or blood vessel formation in tumors <ref> Hogg, PJ. 2002. Biological Regulation through protein disulfide bond cleavage. Redox Report. 7(2), 71-77. </ref> Without the formation of blood vessels to tumors nutrients are limited and tumor growth is limited. Once blood vessels are established growth can rapidly increase. The fact that tumor cells secrete PGK to allow blood vessel formation through the activation of the zymogen plasmin has important implications for understanding its regulation. If the regulation of PGK in tumor cells can be understood, it might be possible to inhibit the overproduction and secretion of PGK to limit angiogenesis in tumors.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-31T17:49:29Z

Shane Harmon: /* Phosphoglycerate Kinase(PGK) */

=Phosphoglycerate Kinase(PGK)=

== PGK in the Glycolysis Cycle ==

Phosphoglycerate kinase is a crucial enzyme in the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphoglycerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/2'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/4'>ATP</scene>. This method for ATP production is known as substrate-level phosphorylation because it produces energy storing ATP molecules without the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously.

{{STRUCTURE_3cin | PDB=1PHP | SCENE= }}

== Structure ==

The overall structure of Phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of approximately 400 amino acids, with a molecular weight of about 45kD. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations: <scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP classification of PGK is alpha and beta, indicating that its <scene name='Shane_Harmon_Sandbox/Scop_classifcation/1'>secondary structure</scene>is composed of roughly equal numbers alpha and beta sheets.

== Reaction Mechanism ==

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. The N-terminal domain has a basic region where the 1,3-Biphosphoglycerate and 3-phosphoglycerate bind while the C-terminal domain has the binding sites for the nucleotide substrates, ADP and ATP. Upon binding of both substrate molecules at the active sites, the protein’s conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. This hydrophobic chamber is necessary to prevent ATP hydrolysis <ref> Auerbach, Gunter et al. 1997. Closed Structure of phosphoglycerate kinase from Thermotoga maritima reveals the catalytic mechanism and determinants of thermal stability. Structure. 5:1475-1483.</ref> The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Arg_and_asp/2'>Arg 62 and Asp 200</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> Recent research indicates that the mechanism for closure of the two domains is a series of hydrogen bond interactions that occur upon binding of the substrates on both domains <ref>Vas, M, Varga, A et al. 2010. Insight into the Mechanism of of Domain Movements and their Role in Enzyme Function: Example of 3-Phosphoglycerate kinase. Current Protein and Peptide Science. Jan 21, 2010. (Epub ahead of publication).</ref>

The mechanism of catalysis has not been fully established because the PGK/1-3biphophoglycerate complex is highly unstable; however, it is thought that the mechanism is similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes the C1 phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref>

The general mechanism is a single displacement Sn2 reaction in which the ADP-B-phosphate oxygen atom initiates nucleophilic attack on the 1-phosphate group of 1-3biphosphoglycerate. Thus, the phosphoryl group is transferred directly via a charged transition state. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form <scene name='Shane_Harmon_Sandbox/Atp/5'>hydrogen bonds</scene> with the enzyme. The 3 hydrogen bonds of ATP are favored over the 2 hydrogen bonds of ADP.

[[Image:PGKmechanism.jpg]]

Two specific residues known to be necessary for catalysis are <scene name='Shane_Harmon_Sandbox/197_and_38/2'>Lys 197 and Arg 36</scene>. Lys 197 secures 1,3-biphosphoblycerate in the closed conformation, and it has been proposed that the transition state intermediary is stabilized by the highly conserved Lys 197 as it transfers the phosphate group. Additionally, it has been shown that Arg 38 is also necessary for catalytic function. Arg 36 has been shown to stabilize a water molecule in the closed conformation and may form a hydrogen bond with the ATP product.

== Kinetics ==

Given that Phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-Hofstee plots curve upward. One possible explanation for the non-linearity, negative cooperativity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 </ref>

== Regulation ==

The role of PGK in glycolysis is very important for the production of ATP through substrate level phosphorylation. Regulation of this protein is thereby controlled by ATP or energy level of the cell. Recent research in frogs which can withstand freezing temperatures indicates that PGK is up regulated by the cold and more specifically by low levels of oxygen <ref> Shaobo, Wu et al. 2009. PGK1 expression responds to freezing, anoxia, and dehydration stresses in freeze tolerant wood frog, Rana sylvatica. Journal of Experimental Zoology. 311, 57-67 </ref> This makes logical sense as when oxygen is low, oxidative phosphorylation cannot occur. Thus without oxygen and oxidative phosphorylation, ATP levels begin to drop. In response to decreased ATP, PGK is up regulated to increase the amount of ATP produced by substrate level phosphorylation. It could therefore also be expected that ADP might act to inhibit or down regulated PGK expression.

Recent study of PGK has revolved around its function in tumor formation and growth. It has been shown that in addition to catalyzing its normal reaction of 1,3-Biphosphoglycerate and ADP to ATP and 3-Phosphoglycerate, PGK can also function to cleave disulfide bonds. Specifically, the review of sulfide bond cleavage indicates PGK has been shown to cleave disulfide bonds in the protein zymogen plasmin to produce the active form of the protein. The active form of plasmin is responsible for angiogenesis or blood vessel formation in tumors <ref> Hogg, PJ. 2002. Biological Regulation through protein disulfide bond cleavage. Redox Report. 7(2), 71-77. </ref> Without the formation of blood vessels to tumors nutrients are limited and tumor growth is limited. Once blood vessels are established growth can rapidly increase. The fact that tumor cells secrete PGK to allow blood vessel formation through the activation of the zymogen plasmin has important implications for understanding its regulation. If the regulation of PGK in tumor cells can be understood, it might be possible to inhibit the overproduction and secretion of PGK to limit angiogenesis in tumors.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-31T17:19:59Z

Shane Harmon: /* Phosphoglycerate Kinase(PGK) */

=Phosphoglycerate Kinase(PGK)=

== PGK in the Glycolysis Cycle ==

Phosphoglycerate kinase is a crucial enzyme in the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphoglycerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/2'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/4'>ATP</scene>. This method for ATP production is known as substrate-level phosphorylation because it produces energy storing ATP molecules without the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously.

{{STRUCTURE_3cin | PDB=1PHP | SCENE= }}

== Structure ==

The overall structure of Phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of approximately 400 amino acids, with a molecular weight of about 45kD. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations: <scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP classification of PGK is alpha and beta, indicating that its <scene name='Shane_Harmon_Sandbox/Scop_classifcation/1'>secondary structure</scene>is composed of roughly equal numbers alpha and beta sheets.

== Reaction Mechanism ==

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. The N-terminal domain has a basic region where the 1,3-Biphosphoglycerate and 3-phosphoglycerate bind while the C-terminal domain has the binding sites for the nucleotide substrates, ADP and ATP. Upon binding of both substrate molecules at the active sites, the protein’s conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. This hydrophobic chamber is necessary to prevent ATP hydrolysis <ref> Auerbach, Gunter et al. 1997. Closed Structure of phosphoglycerate kinase from Thermotoga maritima reveals the catalytic mechanism and determinants of thermal stability. Structure. 5:1475-1483.</ref> The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Arg_and_asp/2'>Arg 62 and Asp 200</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> Recent research indicates that the mechanism for closure of the two domains is a series of hydrogen bond interactions that occur upon binding of the substrates on both domains <ref>Vas, M, Varga, A et al. 2010. Insight into the Mechanism of of Domain Movements and their Role in Enzyme Function: Example of 3-Phosphoglycerate kinase. Current Protein and Peptide Science. Jan 21, 2010. (Epub ahead of publication).</ref>

The mechanism of catalysis has not been fully established because the PGK/1-3biphophoglycerate complex is highly unstable; however, it is thought that the mechanism is similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes the C1 phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref>

The general mechanism is a single displacement Sn2 reaction in which the ADP-B-phosphate oxygen atom initiates nucleophilic attack on the 1-phosphate group of 1-3biphosphoglycerate. Thus, the phosphoryl group is transferred directly via a charged transition state. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form <scene name='Shane_Harmon_Sandbox/Atp/5'>hydrogen bonds</scene> with the enzyme. The 3 hydrogen bonds of ATP are favored over the 2 hydrogen bonds of ADP.

Two specific residues known to be necessary for catalysis are <scene name='Shane_Harmon_Sandbox/197_and_38/2'>Lys 197 and Arg 36</scene>. Lys 197 secures 1,3-biphosphoblycerate in the closed conformation, and it has been proposed that the transition state intermediary is stabilized by the highly conserved Lys 197 as it transfers the phosphate group. Additionally, it has been shown that Arg 38 is also necessary for catalytic function. Arg 36 has been shown to stabilize a water molecule in the closed conformation and may form a hydrogen bond with the ATP product.

== Kinetics ==

Given that Phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-Hofstee plots curve upward. One possible explanation for the non-linearity, negative cooperativity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 </ref>

== Regulation ==

The role of PGK in glycolysis is very important for the production of ATP through substrate level phosphorylation. Regulation of this protein is thereby controlled by ATP or energy level of the cell. Recent research in frogs which can withstand freezing temperatures indicates that PGK is up regulated by the cold and more specifically by low levels of oxygen <ref> Shaobo, Wu et al. 2009. PGK1 expression responds to freezing, anoxia, and dehydration stresses in freeze tolerant wood frog, Rana sylvatica. Journal of Experimental Zoology. 311, 57-67 </ref> This makes logical sense as when oxygen is low, oxidative phosphorylation cannot occur. Thus without oxygen and oxidative phosphorylation, ATP levels begin to drop. In response to decreased ATP, PGK is up regulated to increase the amount of ATP produced by substrate level phosphorylation. It could therefore also be expected that ADP might act to inhibit or down regulated PGK expression.

Recent study of PGK has revolved around its function in tumor formation and growth. It has been shown that in addition to catalyzing its normal reaction of 1,3-Biphosphoglycerate and ADP to ATP and 3-Phosphoglycerate, PGK can also function to cleave disulfide bonds. Specifically, the review of sulfide bond cleavage indicates PGK has been shown to cleave disulfide bonds in the protein zymogen plasmin to produce the active form of the protein. The active form of plasmin is responsible for angiogenesis or blood vessel formation in tumors <ref> Hogg, PJ. 2002. Biological Regulation through protein disulfide bond cleavage. Redox Report. 7(2), 71-77. </ref> Without the formation of blood vessels to tumors nutrients are limited and tumor growth is limited. Once blood vessels are established growth can rapidly increase. The fact that tumor cells secrete PGK to allow blood vessel formation through the activation of the zymogen plasmin has important implications for understanding its regulation. If the regulation of PGK in tumor cells can be understood, it might be possible to inhibit the overproduction and secretion of PGK to limit angiogenesis in tumors.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-31T01:53:31Z

Shane Harmon: /* Phosphoglycerate Kinase(PGK) */

=Phosphoglycerate Kinase(PGK)=

== PGK in the Glycolysis Cycle ==

Phosphoglycerate kinase is a crucial enzyme in the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphoglycerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/2'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/4'>ATP</scene>. This method for ATP production is known as substrate-level phosphorylation because it produces energy storing ATP molecules without the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously.

{{STRUCTURE_3cin | PDB=1PHP | SCENE= }}

== Structure ==

The overall structure of Phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of approximately 400 amino acids, with a molecular weight of about 45kD. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations: <scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP classification of PGK is alpha and beta, indicating that its <scene name='Shane_Harmon_Sandbox/Scop_classifcation/1'>secondary structure</scene>is composed of roughly equal numbers alpha and beta sheets.

== Reaction Mechanism ==

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. The N-terminal domain has a basic region where the 1,3-Biphosphoglycerate and 3-phosphoglycerate bind while the C-terminal domain has the binding sites for the nucleotide substrates, ADP and ATP. Upon binding of both substrate molecules at the active sites, the protein’s conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. This hydrophobic chamber is necessary to prevent ATP hydrolysis <ref> Auerbach, Gunter et al. 1997. Closed Structure of phosphoglycerate kinase from Thermotoga maritima reveals the catalytic mechanism and determinants of thermal stability. Structure. 5:1475-1483.</ref> The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Arg_and_asp/2'>Arg 62 and Asp 200</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> Recent research indicates that the mechanism for closure of the two domains is a series of hydrogen bond interactions that occur upon binding of the substrates on both domains <ref>Vas, M, Varga, A et al. 2010. Insight into the Mechanism of of Domain Movements and their Role in Enzyme Function: Example of 3-Phosphoglycerate kinase. Current Protein and Peptide Science. Jan 21, 2010. (Epub ahead of publication).</ref>

The mechanism of catalysis has not been fully established because the PGK/1-3biphophoglycerate complex is highly unstable; however, it is thought that the mechanism is similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes the C1 phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref>

The general mechanism is a single displacement Sn2 reaction in which the ADP-B-phosphate oxygen atom initiates nucleophilic attack on the 1-phosphate group of 1-3biphosphoglycerate. Thus, the phosphoryl group is transferred directly via a charged transition state. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form <scene name='Shane_Harmon_Sandbox/Atp/5'>hydrogen bonds</scene> with the enzyme. The 3 hydrogen bonds of ATP are favored over the 2 hydrogen bonds of ADP.

Two specific residues known to be necessary for catalysis are <scene name='Shane_Harmon_Sandbox/197_and_38/2'>Lys 197 and Arg 36</scene>. It has been proposed that the transition state intermediary is stabilized by the highly conserved positive Lys 197 as it transfers the phosphate group. Additionally, it has been shown that Arg 38 is also necessary for catalytic function. Arg 36 has been shown to stabilize a water molecule in the closed conformation and may form a hydrogen bond with the ATP product.

== Kinetics ==

Given that Phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-Hofstee plots curve upward. One possible explanation for the non-linearity, negative cooperativity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 </ref>

== Regulation ==

The role of PGK in glycolysis is very important for the production of ATP through substrate level phosphorylation. Regulation of this protein is thereby controlled by ATP or energy level of the cell. Recent research in frogs which can withstand freezing temperatures indicates that PGK is up regulated by the cold and more specifically by low levels of oxygen <ref> Shaobo, Wu et al. 2009. PGK1 expression responds to freezing, anoxia, and dehydration stresses in freeze tolerant wood frog, Rana sylvatica. Journal of Experimental Zoology. 311, 57-67 </ref> This makes logical sense as when oxygen is low, oxidative phosphorylation cannot occur. Thus without oxygen and oxidative phosphorylation, ATP levels begin to drop. In response to decreased ATP, PGK is up regulated to increase the amount of ATP produced by substrate level phosphorylation. It could therefore also be expected that ADP might act to inhibit or down regulated PGK expression.

Recent study of PGK has revolved around its function in tumor formation and growth. It has been shown that in addition to catalyzing its normal reaction of 1,3-Biphosphoglycerate and ADP to ATP and 3-Phosphoglycerate, PGK can also function to cleave disulfide bonds. Specifically, the review of sulfide bond cleavage indicates PGK has been shown to cleave disulfide bonds in the protein zymogen plasmin to produce the active form of the protein. The active form of plasmin is responsible for angiogenesis or blood vessel formation in tumors <ref> Hogg, PJ. 2002. Biological Regulation through protein disulfide bond cleavage. Redox Report. 7(2), 71-77. </ref> Without the formation of blood vessels to tumors nutrients are limited and tumor growth is limited. Once blood vessels are established growth can rapidly increase. The fact that tumor cells secrete PGK to allow blood vessel formation through the activation of the zymogen plasmin has important implications for understanding its regulation. If the regulation of PGK in tumor cells can be understood, it might be possible to inhibit the overproduction and secretion of PGK to limit angiogenesis in tumors.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-31T01:50:05Z

Shane Harmon: /* Phosphoglycerate Kinase(PGK) */

=Phosphoglycerate Kinase(PGK)=

== PGK in the Glycolysis Cycle ==

Phosphoglycerate kinase is a crucial enzyme in the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphoglycerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/2'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/4'>ATP</scene>. This method for ATP production is known as substrate-level phosphorylation because it produces energy storing ATP molecules without the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously.

{{STRUCTURE_3cin | PDB=1PHP | SCENE= }}

== Structure ==

The overall structure of Phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of approximately 400 amino acids, with a molecular weight of about 45kD. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations: <scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP classification of PGK is alpha and beta, indicating that its <scene name='Shane_Harmon_Sandbox/Scop_classifcation/1'>secondary structure</scene>is composed of roughly equal numbers alpha and beta sheets.

== Reaction Mechanism ==

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. The N-terminal domain has a basic region where the 1,3-Biphosphoglycerate and 3-phosphoglycerate bind while the C-terminal domain has the binding sites for the nucleotide substrates, ADP and ATP. Upon binding of both substrate molecules at the active sites, the protein’s conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. This hydrophobic chamber is necessary to prevent ATP hydrolysis <ref> Auerbach, Gunter et al. 1997. Closed Structure of phosphoglycerate kinase from Thermotoga maritima reveals the catalytic mechanism and determinants of thermal stability. Structure. 5:1475-1483.</ref> The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Arg_and_asp/2'>Arg 62 and Asp 200</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> Recent research indicates that the mechanism for closure of the two domains is a series of hydrogen bond interactions that occur upon binding of the substrates on both domains <ref>Vas, M, Varga, A et al. 2010. Insight into the Mechanism of of Domain Movements and their Role in Enzyme Function: Example of 3-Phosphoglycerate kinase. Current Protein and Peptide Science. Jan 21, 2010. (Epub ahead of publication).</ref>

The mechanism of catalysis has not been fully established because the PGK/1-3biphophoglycerate complex is highly unstable; however, it is thought that the mechanism is similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes the C1 phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref>

The general mechanism is a single displacement Sn2 reaction in which the ADP-B-phosphate oxygen atom initiates nucleophilic attack on the 1-phosphate group of 1-3biphosphoglycerate. Thus, the phosphoryl group is transferred directly via a charged transition state. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form <scene name='Shane_Harmon_Sandbox/Atp/5'>hydrogen bonds</scene> with the enzyme. The 3 hydrogen bonds of ATP are favored over the 2 hydrogen bonds of ADP.

Two specific residues known to be necessary for catalysis are <scene name='Shane_Harmon_Sandbox/197_and_38/2'>Lys 197 and Arg 36</scene>. It has been proposed that the transition state intermediary is stabilized by the highly conserved positive Lys 197 as it transfers the phosphate group. Additionally, it has been shown that Arg 38 is also necessary for catalytic function. Arg 36 has been shown to stabilize a water molecule in the closed conformation.

== Kinetics ==

Given that Phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-Hofstee plots curve upward. One possible explanation for the non-linearity, negative cooperativity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 </ref>

== Regulation ==

The role of PGK in glycolysis is very important for the production of ATP through substrate level phosphorylation. Regulation of this protein is thereby controlled by ATP or energy level of the cell. Recent research in frogs which can withstand freezing temperatures indicates that PGK is up regulated by the cold and more specifically by low levels of oxygen <ref> Shaobo, Wu et al. 2009. PGK1 expression responds to freezing, anoxia, and dehydration stresses in freeze tolerant wood frog, Rana sylvatica. Journal of Experimental Zoology. 311, 57-67 </ref> This makes logical sense as when oxygen is low, oxidative phosphorylation cannot occur. Thus without oxygen and oxidative phosphorylation, ATP levels begin to drop. In response to decreased ATP, PGK is up regulated to increase the amount of ATP produced by substrate level phosphorylation. It could therefore also be expected that ADP might act to inhibit or down regulated PGK expression.

Recent study of PGK has revolved around its function in tumor formation and growth. It has been shown that in addition to catalyzing its normal reaction of 1,3-Biphosphoglycerate and ADP to ATP and 3-Phosphoglycerate, PGK can also function to cleave disulfide bonds. Specifically, the review of sulfide bond cleavage indicates PGK has been shown to cleave disulfide bonds in the protein zymogen plasmin to produce the active form of the protein. The active form of plasmin is responsible for angiogenesis or blood vessel formation in tumors <ref> Hogg, PJ. 2002. Biological Regulation through protein disulfide bond cleavage. Redox Report. 7(2), 71-77. </ref> Without the formation of blood vessels to tumors nutrients are limited and tumor growth is limited. Once blood vessels are established growth can rapidly increase. The fact that tumor cells secrete PGK to allow blood vessel formation through the activation of the zymogen plasmin has important implications for understanding its regulation. If the regulation of PGK in tumor cells can be understood, it might be possible to inhibit the overproduction and secretion of PGK to limit angiogenesis in tumors.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-31T01:18:44Z

Shane Harmon: /* Phosphoglycerate Kinase(PGK) */

=Phosphoglycerate Kinase(PGK)=

== PGK in the Glycolysis Cycle ==

Phosphoglycerate kinase is a crucial enzyme in the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphoglycerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/2'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/4'>ATP</scene>. This method for ATP production is known as substrate-level phosphorylation because it produces energy storing ATP molecules without the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously.

{{STRUCTURE_3cin | PDB=1PHP | SCENE= }}

== Structure ==

The overall structure of Phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of approximately 400 amino acids, with a molecular weight of about 45kD. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations: <scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP classification of PGK is alpha and beta, indicating that its <scene name='Shane_Harmon_Sandbox/Scop_classifcation/1'>secondary structure</scene>is composed of roughly equal numbers alpha and beta sheets.

== Reaction Mechanism ==

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. The N-terminal domain has a basic region where the 1,3-Biphosphoglycerate and 3-phosphoglycerate bind while the C-terminal domain has the binding sites for the nucleotide substrates, ADP and ATP. Upon binding of both substrate molecules at the active sites, the protein’s conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. This hydrophobic chamber is necessary to prevent ATP hydrolysis <ref> Auerbach, Gunter et al. 1997. Closed Structure of phosphoglycerate kinase from Thermotoga maritima reveals the catalytic mechanism and determinants of thermal stability. Structure. 5:1475-1483.</ref> The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Arg_and_asp/2'>Arg 62 and Asp 200</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> Recent research indicates that the mechanism for closure of the two domains is a series of hydrogen bond interactions that occur upon binding of the substrates on both domains <ref>Vas, M, Varga, A et al. 2010. Insight into the Mechanism of of Domain Movements and their Role in Enzyme Function: Example of 3-Phosphoglycerate kinase. Current Protein and Peptide Science. Jan 21, 2010. (Epub ahead of publication).</ref>

The mechanism of catalysis has not been fully established because the PGK/1-3biphophoglycerate complex is highly unstable; however, it is thought that the mechanism is similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes the C1 phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref>

The general mechanism is a single displacement Sn2 reaction in which the ADP-B-phosphate oxygen atom initiates nucleophilic attack on the 1-phosphate group of 1-3biphosphoglycerate. Thus, the phosphoryl group is transferred directly via a charged transition state. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form <scene name='Shane_Harmon_Sandbox/Atp/5'>hydrogen bonds</scene> with the enzyme. The 3 hydrogen bonds of ATP are favored over the 2 hydrogen bonds of ADP.

It has been proposed that the transition state intermediary is stabilized by the highly conserved positive Lys 197 as it transfers the phosphate group. Additionally, it has been shown that Arg 38(36) is also necessary for catalytic function. Arg 38(36) has been shown to stabilize a water molecule which is……..

== Kinetics ==

Given that Phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-Hofstee plots curve upward. One possible explanation for the non-linearity, negative cooperativity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 </ref>

== Regulation ==

The role of PGK in glycolysis is very important for the production of ATP through substrate level phosphorylation. Regulation of this protein is thereby controlled by ATP or energy level of the cell. Recent research in frogs which can withstand freezing temperatures indicates that PGK is up regulated by the cold and more specifically by low levels of oxygen <ref> Shaobo, Wu et al. 2009. PGK1 expression responds to freezing, anoxia, and dehydration stresses in freeze tolerant wood frog, Rana sylvatica. Journal of Experimental Zoology. 311, 57-67 </ref> This makes logical sense as when oxygen is low, oxidative phosphorylation cannot occur. Thus without oxygen and oxidative phosphorylation, ATP levels begin to drop. In response to decreased ATP, PGK is up regulated to increase the amount of ATP produced by substrate level phosphorylation. It could therefore also be expected that ADP might act to inhibit or down regulated PGK expression.

Recent study of PGK has revolved around its function in tumor formation and growth. It has been shown that in addition to catalyzing its normal reaction of 1,3-Biphosphoglycerate and ADP to ATP and 3-Phosphoglycerate, PGK can also function to cleave disulfide bonds. Specifically, the review of sulfide bond cleavage indicates PGK has been shown to cleave disulfide bonds in the protein zymogen plasmin to produce the active form of the protein. The active form of plasmin is responsible for angiogenesis or blood vessel formation in tumors <ref> Hogg, PJ. 2002. Biological Regulation through protein disulfide bond cleavage. Redox Report. 7(2), 71-77. </ref> Without the formation of blood vessels to tumors nutrients are limited and tumor growth is limited. Once blood vessels are established growth can rapidly increase. The fact that tumor cells secrete PGK to allow blood vessel formation through the activation of the zymogen plasmin has important implications for understanding its regulation. If the regulation of PGK in tumor cells can be understood, it might be possible to inhibit the overproduction and secretion of PGK to limit angiogenesis in tumors.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-31T01:05:25Z

Shane Harmon: /* Phosphoglycerate Kinase(PGK) */

=Phosphoglycerate Kinase(PGK)=

== PGK in the Glycolysis Cycle ==

Phosphoglycerate kinase is a crucial enzyme in the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphoglycerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/2'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/4'>ATP</scene>. This method for ATP production is known as substrate-level phosphorylation because it produces energy storing ATP molecules without the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously.

{{STRUCTURE_3cin | PDB=1PHP | SCENE= }}

== Structure ==

The overall structure of Phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of approximately 400 amino acids, with a molecular weight of about 45kD. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations: <scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP classification of PGK is alpha and beta, indicating that its secondary structure is composed of roughly equal numbers alpha and beta sheets.

== Reaction Mechanism ==

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. The N-terminal domain has a basic region where the 1,3-Biphosphoglycerate and 3-phosphoglycerate bind while the C-terminal domain has the binding sites for the nucleotide substrates, ADP and ATP. Upon binding of both substrate molecules at the active sites, the protein’s conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. This hydrophobic chamber is necessary to prevent ATP hydrolysis <ref> Auerbach, Gunter et al. 1997. Closed Structure of phosphoglycerate kinase from Thermotoga maritima reveals the catalytic mechanism and determinants of thermal stability. Structure. 5:1475-1483.</ref> The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Arg_and_asp/2'>Arg 62 and Asp 200</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> Recent research indicates that the mechanism for closure of the two domains is a series of hydrogen bond interactions that occur upon binding of the substrates on both domains <ref>Vas, M, Varga, A et al. 2010. Insight into the Mechanism of of Domain Movements and their Role in Enzyme Function: Example of 3-Phosphoglycerate kinase. Current Protein and Peptide Science. Jan 21, 2010. (Epub ahead of publication).</ref>

The mechanism of catalysis has not been fully established because the PGK/1-3biphophoglycerate complex is highly unstable; however, it is thought that the mechanism is similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes the C1 phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref>

The general mechanism is a single displacement Sn2 reaction in which the ADP-B-phosphate oxygen atom initiates nucleophilic attack on the 1-phosphate group of 1-3biphosphoglycerate. Thus, the phosphoryl group is transferred directly via a charged transition state. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form <scene name='Shane_Harmon_Sandbox/Atp/5'>hydrogen bonds</scene> with the enzyme. The 3 hydrogen bonds of ATP are favored over the 2 hydrogen bonds of ADP.

It has been proposed that the transition state intermediary is stabilized by the highly conserved positive Lys 197 as it transfers the phosphate group. Additionally, it has been shown that Arg 38(36) is also necessary for catalytic function. Arg 38(36) has been shown to stabilize a water molecule which is……..

== Kinetics ==

Given that Phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-Hofstee plots curve upward. One possible explanation for the non-linearity, negative cooperativity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 </ref>

== Regulation ==

The role of PGK in glycolysis is very important for the production of ATP through substrate level phosphorylation. Regulation of this protein is thereby controlled by ATP or energy level of the cell. Recent research in frogs which can withstand freezing temperatures indicates that PGK is up regulated by the cold and more specifically by low levels of oxygen <ref> Shaobo, Wu et al. 2009. PGK1 expression responds to freezing, anoxia, and dehydration stresses in freeze tolerant wood frog, Rana sylvatica. Journal of Experimental Zoology. 311, 57-67 </ref> This makes logical sense as when oxygen is low, oxidative phosphorylation cannot occur. Thus without oxygen and oxidative phosphorylation, ATP levels begin to drop. In response to decreased ATP, PGK is up regulated to increase the amount of ATP produced by substrate level phosphorylation. It could therefore also be expected that ADP might act to inhibit or down regulated PGK expression.

Recent study of PGK has revolved around its function in tumor formation and growth. It has been shown that in addition to catalyzing its normal reaction of 1,3-Biphosphoglycerate and ADP to ATP and 3-Phosphoglycerate, PGK can also function to cleave disulfide bonds. Specifically, the review of sulfide bond cleavage indicates PGK has been shown to cleave disulfide bonds in the protein zymogen plasmin to produce the active form of the protein. The active form of plasmin is responsible for angiogenesis or blood vessel formation in tumors <ref> Hogg, PJ. 2002. Biological Regulation through protein disulfide bond cleavage. Redox Report. 7(2), 71-77. </ref> Without the formation of blood vessels to tumors nutrients are limited and tumor growth is limited. Once blood vessels are established growth can rapidly increase. The fact that tumor cells secrete PGK to allow blood vessel formation through the activation of the zymogen plasmin has important implications for understanding its regulation. If the regulation of PGK in tumor cells can be understood, it might be possible to inhibit the overproduction and secretion of PGK to limit angiogenesis in tumors.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-31T00:51:48Z

Shane Harmon: /* Phosphoglycerate Kinase(PGK) */

=Phosphoglycerate Kinase(PGK)=

== PGK in the Glycolysis Cycle ==

Phosphoglycerate kinase is a crucial enzyme in the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphoglycerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/2'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/4'>ATP</scene>. This method for ATP production is known as substrate-level phosphorylation because it produces energy storing ATP molecules without the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously.

{{STRUCTURE_3cin | PDB=1PHP | SCENE= }}

== Structure ==

The overall structure of Phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of approximately 400 amino acids, with a molecular weight of about 45kD. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations: <scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP classification of PGK is alpha and beta, indicating that its secondary structure is composed of roughly equal numbers alpha and beta sheets.

== Reaction Mechanism ==

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. The N-terminal domain has a basic region where the 1,3-Biphosphoglycerate and 3-phosphoglycerate bind while the C-terminal domain has the binding sites for the nucleotide substrates, ADP and ATP. Upon binding of both substrate molecules at the active sites, the protein’s conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. This hydrophobic chamber is necessary to prevent ATP hydrolysis <ref> Auerbach, Gunter et al. 1997. Closed Structure of phosphoglycerate kinase from Thermotoga maritima reveals the catalytic mechanism and determinants of thermal stability. Structure. 5:1475-1483.</ref> The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Arg_and_asp/1'>Arg62 and Asp200</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> Recent research indicates that the mechanism for closure of the two domains is a series of hydrogen bond interactions that occur upon binding of the substrates on both domains <ref>Vas, M, Varga, A et al. 2010. Insight into the Mechanism of of Domain Movements and their Role in Enzyme Function: Example of 3-Phosphoglycerate kinase. Current Protein and Peptide Science. Jan 21, 2010. (Epub ahead of publication).</ref>

The mechanism of catalysis has not been fully established because the PGK/1-3biphophoglycerate complex is highly unstable; however, it is thought that the mechanism is similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes the C1 phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref>

The general mechanism is a single displacement Sn2 reaction in which the ADP-B-phosphate oxygen atom initiates nucleophilic attack on the 1-phosphate group of 1-3biphosphoglycerate. Thus, the phosphoryl group is transferred directly via a charged transition state. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form <scene name='Shane_Harmon_Sandbox/Atp/5'>hydrogen bonds</scene> with the enzyme. The 3 hydrogen bonds of ATP are favored over the 2 hydrogen bonds of ADP.

It has been proposed that the transition state intermediary is stabilized by the highly conserved positive Lys 197 as it transfers the phosphate group. Additionally, it has been shown that Arg 38(36) is also necessary for catalytic function. Arg 38(36) has been shown to stabilize a water molecule which is……..

== Kinetics ==

Given that Phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-Hofstee plots curve upward. One possible explanation for the non-linearity, negative cooperativity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 </ref>

== Regulation ==

The role of PGK in glycolysis is very important for the production of ATP through substrate level phosphorylation. Regulation of this protein is thereby controlled by ATP or energy level of the cell. Recent research in frogs which can withstand freezing temperatures indicates that PGK is up regulated by the cold and more specifically by low levels of oxygen <ref> Shaobo, Wu et al. 2009. PGK1 expression responds to freezing, anoxia, and dehydration stresses in freeze tolerant wood frog, Rana sylvatica. Journal of Experimental Zoology. 311, 57-67 </ref> This makes logical sense as when oxygen is low, oxidative phosphorylation cannot occur. Thus without oxygen and oxidative phosphorylation, ATP levels begin to drop. In response to decreased ATP, PGK is up regulated to increase the amount of ATP produced by substrate level phosphorylation. It could therefore also be expected that ADP might act to inhibit or down regulated PGK expression.

Recent study of PGK has revolved around its function in tumor formation and growth. It has been shown that in addition to catalyzing its normal reaction of 1,3-Biphosphoglycerate and ADP to ATP and 3-Phosphoglycerate, PGK can also function to cleave disulfide bonds. Specifically, the review of sulfide bond cleavage indicates PGK has been shown to cleave disulfide bonds in the protein zymogen plasmin to produce the active form of the protein. The active form of plasmin is responsible for angiogenesis or blood vessel formation in tumors <ref> Hogg, PJ. 2002. Biological Regulation through protein disulfide bond cleavage. Redox Report. 7(2), 71-77. </ref> Without the formation of blood vessels to tumors nutrients are limited and tumor growth is limited. Once blood vessels are established growth can rapidly increase. The fact that tumor cells secrete PGK to allow blood vessel formation through the activation of the zymogen plasmin has important implications for understanding its regulation. If the regulation of PGK in tumor cells can be understood, it might be possible to inhibit the overproduction and secretion of PGK to limit angiogenesis in tumors.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-30T23:57:34Z

Shane Harmon: /* Phosphoglycerate Kinase(PGK) */

=Phosphoglycerate Kinase(PGK)=

== PGK in the Glycolysis Cycle ==

Phosphoglycerate kinase is a crucial enzyme in the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphoglycerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/2'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/4'>ATP</scene>. This method for ATP production is known as substrate-level phosphorylation because it produces energy storing ATP molecules without the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously.

{{STRUCTURE_3cin | PDB=1PHP | SCENE= }}

== Structure ==

The overall structure of Phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of approximately 400 amino acids, with a molecular weight of about 45kD. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations: <scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP classification of PGK is alpha and beta, indicating that its secondary structure is composed of roughly equal numbers alpha and beta sheets.

== Reaction Mechanism ==

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. The N-terminal domain has a basic region where the 1,3-Biphosphoglycerate and 3-phosphoglycerate bind while the C-terminal domain has the binding sites for the nucleotide substrates, ADP and ATP. Upon binding of both substrate molecules at the active sites, the protein’s conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. This hydrophobic chamber is necessary to prevent ATP hydrolysis <ref> Auerbach, Gunter et al. 1997. Closed Structure of phosphoglycerate kinase from Thermotoga maritima reveals the catalytic mechanism and determinants of thermal stability. Structure. 5:1475-1483.</ref> The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Arg_and_asp/1'>Arg62 and Asp200</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> Recent research indicates that the mechanism for closure of the two domains is a series of hydrogen bond interactions that occur upon binding of the substrates on both domains <ref>Vas, M, Varga, A et al. 2010. Insight into the Mechanism of of Domain Movements and their Role in Enzyme Function: Example of 3-Phosphoglycerate kinase. Current Protein and Peptide Science. Jan 21, 2010. (Epub ahead of publication).</ref>

The mechanism of catalysis has not been fully established because the PGK/1-3biphophoglycerate complex is highly unstable; however, it is thought that the mechanism is similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes the C1 phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref>

The general mechanism is a single displacement Sn2 reaction in which the ADP-B-phosphate oxygen atom initiates nucleophilic attack on the 1-phosphate group of 1-3biphosphoglycerate. Thus, the phosphoryl group is transferred directly via a charged transition state. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form <scene name='Shane_Harmon_Sandbox/Atp/3'>hydrogen bonds</scene> with the enzyme. The 3 hydrogen bonds of ATP are favored over the 2 hydrogen bonds of ADP.

It has been proposed that the transition state intermediary is stabilized by the highly conserved positive Lys 197 as it transfers the phosphate group. Additionally, it has been shown that Arg 38(36) is also necessary for catalytic function. Arg 38(36) has been shown to stabilize a water molecule which is……..

== Kinetics ==

Given that Phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-Hofstee plots curve upward. One possible explanation for the non-linearity, negative cooperativity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 </ref>

== Regulation ==

The role of PGK in glycolysis is very important for the production of ATP through substrate level phosphorylation. Regulation of this protein is thereby controlled by ATP or energy level of the cell. Recent research in frogs which can withstand freezing temperatures indicates that PGK is up regulated by the cold and more specifically by low levels of oxygen <ref> Shaobo, Wu et al. 2009. PGK1 expression responds to freezing, anoxia, and dehydration stresses in freeze tolerant wood frog, Rana sylvatica. Journal of Experimental Zoology. 311, 57-67 </ref> This makes logical sense as when oxygen is low, oxidative phosphorylation cannot occur. Thus without oxygen and oxidative phosphorylation, ATP levels begin to drop. In response to decreased ATP, PGK is up regulated to increase the amount of ATP produced by substrate level phosphorylation. It could therefore also be expected that ADP might act to inhibit or down regulated PGK expression.

Recent study of PGK has revolved around its function in tumor formation and growth. It has been shown that in addition to catalyzing its normal reaction of 1,3-Biphosphoglycerate and ADP to ATP and 3-Phosphoglycerate, PGK can also function to cleave disulfide bonds. Specifically, the review of sulfide bond cleavage indicates PGK has been shown to cleave disulfide bonds in the protein zymogen plasmin to produce the active form of the protein. The active form of plasmin is responsible for angiogenesis or blood vessel formation in tumors <ref> Hogg, PJ. 2002. Biological Regulation through protein disulfide bond cleavage. Redox Report. 7(2), 71-77. </ref> Without the formation of blood vessels to tumors nutrients are limited and tumor growth is limited. Once blood vessels are established growth can rapidly increase. The fact that tumor cells secrete PGK to allow blood vessel formation through the activation of the zymogen plasmin has important implications for understanding its regulation. If the regulation of PGK in tumor cells can be understood, it might be possible to inhibit the overproduction and secretion of PGK to limit angiogenesis in tumors.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-30T15:11:22Z

Shane Harmon: /* Phosphoglycerate Kinase(PGK) */

=Phosphoglycerate Kinase(PGK)=

== PGK in the Glycolysis Cycle ==

Phosphoglycerate kinase is a crucial enzyme in the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphoglycerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/2'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/4'>ATP</scene>. This method for ATP production is known as substrate-level phosphorylation because it produces energy storing ATP molecules without the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously. {{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

{{STRUCTURE_3cin | PDB=1PHP | SCENE= }}

== Structure ==

The overall structure of Phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of approximately 400 amino acids, with a molecular weight of about 45kD. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations: <scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP classification of PGK is alpha and beta, indicating that its secondary structure is composed of roughly equal numbers alpha and beta sheets.

== Reaction Mechanism ==

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. The N-terminal domain has a basic region where the 1,3-Biphosphoglycerate and 3-phosphoglycerate bind while the C-terminal domain has the binding sites for the nucleotide substrates, ADP and ATP. Upon binding of both substrate molecules at the active sites, the protein’s conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. This hydrophobic chamber is necessary to prevent ATP hydrolysis <ref> Auerbach, Gunter et al. 1997. Closed Structure of phosphoglycerate kinase from Thermotoga maritima reveals the catalytic mechanism and determinants of thermal stability. Structure. 5:1475-1483.</ref> The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Arg_and_asp/1'>Arg62 and Asp200</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> Recent research indicates that the mechanism for closure of the two domains is a series of hydrogen bond interactions that occur upon binding of the substrates on both domains <ref>Vas, M, Varga, A et al. 2010. Insight into the Mechanism of of Domain Movements and their Role in Enzyme Function: Example of 3-Phosphoglycerate kinase. Current Protein and Peptide Science. Jan 21, 2010. (Epub ahead of publication).</ref>

The mechanism of catalysis has not been fully established because the PGK/1-3biphophoglycerate complex is highly unstable; however, it is thought that the mechanism is similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes the C1 phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref>

The general mechanism is a single displacement Sn2 reaction in which the ADP-B-phosphate oxygen atom initiates nucleophilic attack on the 1-phosphate group of 1-3biphosphoglycerate. Thus, the phosphoryl group is transferred directly via a charged transition state. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form <scene name='Shane_Harmon_Sandbox/Atp/3'>hydrogen bonds</scene> with the enzyme. The 3 hydrogen bonds of ATP are favored over the 2 hydrogen bonds of ADP.

It has been proposed that the transition state intermediary is stabilized by the highly conserved positive Lys 197 as it transfers the phosphate group. Additionally, it has been shown that Arg 38(36) is also necessary for catalytic function. Arg 38(36) has been shown to stabilize a water molecule which is……..

== Kinetics ==

Given that Phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-Hofstee plots curve upward. One possible explanation for the non-linearity, negative cooperativity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 </ref>

== Regulation ==

The role of PGK in glycolysis is very important for the production of ATP through substrate level phosphorylation. Regulation of this protein is thereby controlled by ATP or energy level of the cell. Recent research in frogs which can withstand freezing temperatures indicates that PGK is up regulated by the cold and more specifically by low levels of oxygen <ref> Shaobo, Wu et al. 2009. PGK1 expression responds to freezing, anoxia, and dehydration stresses in freeze tolerant wood frog, Rana sylvatica. Journal of Experimental Zoology. 311, 57-67 </ref> This makes logical sense as when oxygen is low, oxidative phosphorylation cannot occur. Thus without oxygen and oxidative phosphorylation, ATP levels begin to drop. In response to decreased ATP, PGK is up regulated to increase the amount of ATP produced by substrate level phosphorylation. It could therefore also be expected that ADP might act to inhibit or down regulated PGK expression.

Recent study of PGK has revolved around its function in tumor formation and growth. It has been shown that in addition to catalyzing its normal reaction of 1,3-Biphosphoglycerate and ADP to ATP and 3-Phosphoglycerate, PGK can also function to cleave disulfide bonds. Specifically, the review of sulfide bond cleavage indicates PGK has been shown to cleave disulfide bonds in the protein zymogen plasmin to produce the active form of the protein. The active form of plasmin is responsible for angiogenesis or blood vessel formation in tumors <ref> Hogg, PJ. 2002. Biological Regulation through protein disulfide bond cleavage. Redox Report. 7(2), 71-77. </ref> Without the formation of blood vessels to tumors nutrients are limited and tumor growth is limited. Once blood vessels are established growth can rapidly increase. The fact that tumor cells secrete PGK to allow blood vessel formation through the activation of the zymogen plasmin has important implications for understanding its regulation. If the regulation of PGK in tumor cells can be understood, it might be possible to inhibit the overproduction and secretion of PGK to limit angiogenesis in tumors.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-30T14:58:17Z

Shane Harmon: /* Phosphoglycerate Kinase(PGK) */

=Phosphoglycerate Kinase(PGK)=

== PGK in the Glycolysis Cycle ==

Phosphoglycerate kinase is a crucial enzyme in the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphoglycerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/2'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/4'>ATP</scene>. This method for ATP production is known as substrate-level phosphorylation because it produces energy storing ATP molecules without the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously. {{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

{{STRUCTURE_3cin | PDB=1PHP | SCENE= }}

== Structure ==

The overall structure of Phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of approximately 400 amino acids, with a molecular weight of about 45kD. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations: <scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP classification of PGK is alpha and beta, indicating that its secondary structure is composed of roughly equal numbers alpha and beta sheets.

== Reaction Mechanism ==

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. The N-terminal domain has a basic region where the 1,3-Biphosphoglycerate and 3-phosphoglycerate bind while the C-terminal domain has the binding sites for the nucleotide substrates, ADP and ATP. Upon binding of both substrate molecules at the active sites, the protein’s conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. This hydrophobic chamber is necessary to prevent ATP hydrolysis <ref> Auerbach, Gunter et al. 1997. Closed Structure of phosphoglycerate kinase from Thermotoga maritima reveals the catalytic mechanism and determinants of thermal stability. Structure. 5:1475-1483.</ref> The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> Recent research indicates that the mechanism for closure of the two domains is a series of hydrogen bond interactions that occur upon binding of the substrates on both domains <ref>Vas, M, Varga, A et al. 2010. Insight into the Mechanism of of Domain Movements and their Role in Enzyme Function: Example of 3-Phosphoglycerate kinase. Current Protein and Peptide Science. Jan 21, 2010. (Epub ahead of publication).</ref>

The mechanism of catalysis has not been fully established because the PGK/1-3biphophoglycerate complex is highly unstable; however, it is thought that the mechanism is similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes the C1 phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref>

The general mechanism is a single displacement Sn2 reaction in which the ADP-B-phosphate oxygen atom initiates nucleophilic attack on the 1-phosphate group of 1-3biphosphoglycerate. Thus, the phosphoryl group is transferred directly via a charged transition state. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form <scene name='Shane_Harmon_Sandbox/Atp/3'>hydrogen bonds</scene> with the enzyme. The 3 hydrogen bonds of ATP are favored over the 2 hydrogen bonds of ADP.

It has been proposed that the transition state intermediary is stabilized by the highly conserved positive Lys 197 as it transfers the phosphate group. Additionally, it has been shown that Arg 38(36) is also necessary for catalytic function. Arg 38(36) has been shown to stabilize a water molecule which is……..

== Kinetics ==

Given that Phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-Hofstee plots curve upward. One possible explanation for the non-linearity, negative cooperativity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 </ref>

== Regulation ==

The role of PGK in glycolysis is very important for the production of ATP through substrate level phosphorylation. Regulation of this protein is thereby controlled by ATP or energy level of the cell. Recent research in frogs which can withstand freezing temperatures indicates that PGK is up regulated by the cold and more specifically by low levels of oxygen <ref> Shaobo, Wu et al. 2009. PGK1 expression responds to freezing, anoxia, and dehydration stresses in freeze tolerant wood frog, Rana sylvatica. Journal of Experimental Zoology. 311, 57-67 </ref> This makes logical sense as when oxygen is low, oxidative phosphorylation cannot occur. Thus without oxygen and oxidative phosphorylation, ATP levels begin to drop. In response to decreased ATP, PGK is up regulated to increase the amount of ATP produced by substrate level phosphorylation. It could therefore also be expected that ADP might act to inhibit or down regulated PGK expression.

Recent study of PGK has revolved around its function in tumor formation and growth. It has been shown that in addition to catalyzing its normal reaction of 1,3-Biphosphoglycerate and ADP to ATP and 3-Phosphoglycerate, PGK can also function to cleave disulfide bonds. Specifically, the review of sulfide bond cleavage indicates PGK has been shown to cleave disulfide bonds in the protein zymogen plasmin to produce the active form of the protein. The active form of plasmin is responsible for angiogenesis or blood vessel formation in tumors <ref> Hogg, PJ. 2002. Biological Regulation through protein disulfide bond cleavage. Redox Report. 7(2), 71-77. </ref> Without the formation of blood vessels to tumors nutrients are limited and tumor growth is limited. Once blood vessels are established growth can rapidly increase. The fact that tumor cells secrete PGK to allow blood vessel formation through the activation of the zymogen plasmin has important implications for understanding its regulation. If the regulation of PGK in tumor cells can be understood, it might be possible to inhibit the overproduction and secretion of PGK to limit angiogenesis in tumors.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-30T14:56:53Z

Shane Harmon: /* Phosphoglycerate Kinase(PGK) */

=Phosphoglycerate Kinase(PGK)=

== PGK in the Glycolysis Cycle ==

Phosphoglycerate kinase is a crucial enzyme in the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphoglycerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/2'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/4'>ATP</scene>. This method for ATP production is known as substrate-level phosphorylation because it produces energy storing ATP molecules without the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously. {{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

{{STRUCTURE_3cin | PDB=1PHP | SCENE= }}

== Structure ==

The overall structure of Phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of approximately 400 amino acids, with a molecular weight of about 45kD. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations: <scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP classification of PGK is alpha and beta, indicating that its secondary structure is composed of roughly equal numbers alpha and beta sheets.

== Reaction Mechanism ==

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. The N-terminal domain has a basic region where the 1,3-Biphosphoglycerate and 3-phosphoglycerate bind while the C-terminal domain has the binding sites for the nucleotide substrates, ADP and ATP. Upon binding of both substrate molecules at the active sites, the protein’s conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. This hydrophobic chamber is necessary to prevent ATP hydrolysis <ref> Auerbach, Gunter et al. 1997. Closed Structure of phosphoglycerate kinase from Thermotoga maritima reveals the catalytic mechanism and determinants of thermal stability. Structure. 5:1475-1483.</ref> The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> Recent research indicates that the mechanism for closure of the two domains is a series of hydrogen bond interactions that occur upon binding of the substrates on both domains <ref>Vas, M, Varga, A et al. 2010. Insight into the Mechanism of of Domain Movements and their Role in Enzyme Function: Example of 3-Phosphoglycerate kinase. Current Protein and Peptide Science. Jan 21, 2010. (Epub ahead of publication).</ref>

The mechanism of catalysis has not been fully established because the PGK/1-3biphophoglycerate complex is highly unstable; however, it is thought that the mechanism is similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes the C1 phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref>

The general mechanism is a single displacement Sn2 reaction in which the ADP-B-phosphate oxygen atom initiates nucleophilic attack on the 1-phosphate group of 1-3biphosphoglycerate. Thus, the phosphoryl group is transferred directly via a charged transition state. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form <scene name='Shane_Harmon_Sandbox/Atp/3'>hydrogen bonds</scene> with the enzyme. The 3 hydrogen bonds of ATP are favored over the 2 hydrogen bonds of ADP.

It has been proposed that the transition state intermediary is stabilized by the highly conserved positive Lys 197 as it transfers the phosphate group. Additionally, it has been shown that Arg 38(36) is also necessary for catalytic function. Arg 38(36) has been shown to stabilize a water molecule which is……..

== Kinetics ==

Given that Phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-Hofstee plots curve upward. One possible explanation for the non-linearity, negative cooperativity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 </ref>

== Reaction Mechanism ==

The mechanism of catalysis has not been established but must be similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes a phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref> The negatively charged oxygen of the last phosphate group on ADP nucleophilically attacks a phosphate of 1,3-phosphoglycerate. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form <scene name='Shane_Harmon_Sandbox/Atp/3'>hydrogen bonds</scene> with the enzyme. The 3 hydrogen bonds of ATP is favored over the 2 hydrogen bonds of ADP.

== Regulation ==

The role of PGK in glycolysis is very important for the production of ATP through substrate level phosphorylation. Regulation of this protein is thereby controlled by ATP or energy level of the cell. Recent research in frogs which can withstand freezing temperatures indicates that PGK is up regulated by the cold and more specifically by low levels of oxygen <ref> Shaobo, Wu et al. 2009. PGK1 expression responds to freezing, anoxia, and dehydration stresses in freeze tolerant wood frog, Rana sylvatica. Journal of Experimental Zoology. 311, 57-67 </ref> This makes logical sense as when oxygen is low, oxidative phosphorylation cannot occur. Thus without oxygen and oxidative phosphorylation, ATP levels begin to drop. In response to decreased ATP, PGK is up regulated to increase the amount of ATP produced by substrate level phosphorylation. It could therefore also be expected that ADP might act to inhibit or down regulated PGK expression.

Recent study of PGK has revolved around its function in tumor formation and growth. It has been shown that in addition to catalyzing its normal reaction of 1,3-Biphosphoglycerate and ADP to ATP and 3-Phosphoglycerate, PGK can also function to cleave disulfide bonds. Specifically, the review of sulfide bond cleavage indicates PGK has been shown to cleave disulfide bonds in the protein zymogen plasmin to produce the active form of the protein. The active form of plasmin is responsible for angiogenesis or blood vessel formation in tumors <ref> Hogg, PJ. 2002. Biological Regulation through protein disulfide bond cleavage. Redox Report. 7(2), 71-77. </ref> Without the formation of blood vessels to tumors nutrients are limited and tumor growth is limited. Once blood vessels are established growth can rapidly increase. The fact that tumor cells secrete PGK to allow blood vessel formation through the activation of the zymogen plasmin has important implications for understanding its regulation. If the regulation of PGK in tumor cells can be understood, it might be possible to inhibit the overproduction and secretion of PGK to limit angiogenesis in tumors.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-30T02:21:15Z

Shane Harmon: /* Phosphoglycerate Kinase(PGK) */

=Phosphoglycerate Kinase(PGK)=

== PGK in the Glycolysis Cycle ==

Phosphoglycerate kinase is a crucial enzyme in the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphoglycerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/2'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/4'>ATP</scene>. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously. {{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

{{STRUCTURE_3cin | PDB=1PHP | SCENE= }}

== Structure ==

The overall structure of Phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of XXXXX amino acids. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP clssification of PGK is alpha and beta, indicating that its secondary strucutre is composed of roughly equal numbers alpha and beta sheets.

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrate molecules at the active sites, the proteins conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> Recent research indicates that the mechanism for closure of the two domains is a series of hydrogen bond interactions that occur upon binding of the substrates on both domains <ref>Vas, M, Varga, A et al. 2010. Insight into the Mechanism of of Domain Movements and their Role in Enzyme Function: Example of 3-Phosphoglycerate kinase. Current Protein and Peptide Science. Jan 21, 2010. (Epub ahead of publication).</ref>

== Kinetics ==

Given that Phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-Hofstee plots curve upward. One possible explanation for the non-linearity, negative cooperativity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 </ref>

== Reaction Mechanism ==

The mechanism of catalysis has not been established but must be similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes a phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref> The negatively charged oxygen of the last phosphate group on ADP nucleophilically attacks a phosphate of 1,3-phosphoglycerate. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form <scene name='Shane_Harmon_Sandbox/Atp/3'>hydrogen bonds</scene> with the enzyme. The 3 hydrogen bonds of ATP is favored over the 2 hydrogen bonds of ADP.

== Regulation ==

The role of PGK in glycolysis is very important for the production of ATP through substrate level phosphorylation. Regulation of this protein is thereby controlled by ATP or energy level of the cell. Recent research in frogs which can withstand freezing temperatures indicates that PGK is up regulated by the cold and more specifically by low levels of oxygen <ref> Shaobo, Wu et al. 2009. PGK1 expression responds to freezing, anoxia, and dehydration stresses in freeze tolerant wood frog, Rana sylvatica. Journal of Experimental Zoology. 311, 57-67 </ref> This makes logical sense as when oxygen is low, oxidative phosphorylation cannot occur. Thus without oxygen and oxidative phosphorylation, ATP levels begin to drop. In response to decreased ATP, PGK is up regulated to increase the amount of ATP produced by substrate level phosphorylation. It could therefore also be expected that ADP might act to inhibit or down regulated PGK expression.

Recent study of PGK has revolved around its function in tumor formation and growth. It has been shown that in addition to catalyzing its normal reaction of 1,3-Biphosphoglycerate and ADP to ATP and 3-Phosphoglycerate, PGK can also function to cleave disulfide bonds. Specifically, the review of sulfide bond cleavage indicates PGK has been shown to cleave disulfide bonds in the protein zymogen plasmin to produce the active form of the protein. The active form of plasmin is responsible for angiogenesis or blood vessel formation in tumors <ref> Hogg, PJ. 2002. Biological Regulation through protein disulfide bond cleavage. Redox Report. 7(2), 71-77. </ref> Without the formation of blood vessels to tumors nutrients are limited and tumor growth is limited. Once blood vessels are established growth can rapidly increase. The fact that tumor cells secrete PGK to allow blood vessel formation through the activation of the zymogen plasmin has important implications for understanding its regulation. If the regulation of PGK in tumor cells can be understood, it might be possible to inhibit the overproduction and secretion of PGK to limit angiogenesis in tumors.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-30T01:52:27Z

Shane Harmon: /* Phosphoglycerate Kinase(PGK) */

=Phosphoglycerate Kinase(PGK)=

== PGK in the Glycolysis Cycle ==

Phosphoglycerate kinase is a crucial enzyme in the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphoglycerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/2'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/4'>ATP</scene>. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously. {{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

{{STRUCTURE_3cin | PDB=3C3C | SCENE= }}

== Structure ==

The overall structure of Phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of XXXXX amino acids. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP clssification of PGK is alpha and beta, indicating that its secondary strucutre is composed of roughly equal numbers alpha and beta sheets.

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrate molecules at the active sites, the proteins conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> Recent research indicates that the mechanism for closure of the two domains is a series of hydrogen bond interactions that occur upon binding of the substrates on both domains <ref>Vas, M, Varga, A et al. 2010. Insight into the Mechanism of of Domain Movements and their Role in Enzyme Function: Example of 3-Phosphoglycerate kinase. Current Protein and Peptide Science. Jan 21, 2010. (Epub ahead of publication).</ref>

== Kinetics ==

Given that Phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-Hofstee plots curve upward. One possible explanation for the non-linearity, negative cooperativity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 </ref>

== Reaction Mechanism ==

The mechanism of catalysis has not been established but must be similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes a phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref> The negatively charged oxygen of the last phosphate group on ADP nucleophilically attacks a phosphate of 1,3-phosphoglycerate. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form <scene name='Shane_Harmon_Sandbox/Atp/3'>hydrogen bonds</scene> with the enzyme. The 3 hydrogen bonds of ATP is favored over the 2 hydrogen bonds of ADP.

== Regulation ==

The role of PGK in glycolysis is very important for the production of ATP through substrate level phosphorylation. Regulation of this protein is thereby controlled by ATP or energy level of the cell. Recent research in frogs which can withstand freezing temperatures indicates that PGK is up regulated by the cold and more specifically by low levels of oxygen <ref> Shaobo, Wu et al. 2009. PGK1 expression responds to freezing, anoxia, and dehydration stresses in freeze tolerant wood frog, Rana sylvatica. Journal of Experimental Zoology. 311, 57-67 </ref> This makes logical sense as when oxygen is low, oxidative phosphorylation cannot occur. Thus without oxygen and oxidative phosphorylation, ATP levels begin to drop. In response to decreased ATP, PGK is up regulated to increase the amount of ATP produced by substrate level phosphorylation. It could therefore also be expected that ADP might act to inhibit or down regulated PGK expression.

Recent study of PGK has revolved around its function in tumor formation and growth. It has been shown that in addition to catalyzing its normal reaction of 1,3-Biphosphoglycerate and ADP to ATP and 3-Phosphoglycerate, PGK can also function to cleave disulfide bonds. Specifically, the review of sulfide bond cleavage indicates PGK has been shown to cleave disulfide bonds in the protein zymogen plasmin to produce the active form of the protein. The active form of plasmin is responsible for angiogenesis or blood vessel formation in tumors <ref> Hogg, PJ. 2002. Biological Regulation through protein disulfide bond cleavage. Redox Report. 7(2), 71-77. </ref> Without the formation of blood vessels to tumors nutrients are limited and tumor growth is limited. Once blood vessels are established growth can rapidly increase. The fact that tumor cells secrete PGK to allow blood vessel formation through the activation of the zymogen plasmin has important implications for understanding its regulation. If the regulation of PGK in tumor cells can be understood, it might be possible to inhibit the overproduction and secretion of PGK to limit angiogenesis in tumors.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-30T01:46:30Z

Shane Harmon: /* Mechanism of Phosphoglycerate Kinase(PGK) Catalysis */

=Phosphoglycerate Kinase(PGK)=

== PGK in the Glycolysis Cycle ==

Phosphoglycerate kinase is a crucial enzyme in the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphoglycerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/2'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/4'>ATP</scene>. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously. {{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

{{STRUCTURE_3cin | PDB=3pgk | SCENE= }}

== Structure ==

The overall structure of Phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of XXXXX amino acids. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP clssification of PGK is alpha and beta, indicating that its secondary strucutre is composed of roughly equal numbers alpha and beta sheets.

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrate molecules at the active sites, the proteins conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> Recent research indicates that the mechanism for closure of the two domains is a series of hydrogen bond interactions that occur upon binding of the substrates on both domains <ref>Vas, M, Varga, A et al. 2010. Insight into the Mechanism of of Domain Movements and their Role in Enzyme Function: Example of 3-Phosphoglycerate kinase. Current Protein and Peptide Science. Jan 21, 2010. (Epub ahead of publication).</ref>

== Kinetics ==

Given that Phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-Hofstee plots curve upward. One possible explanation for the non-linearity, negative cooperativity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 </ref>

== Reaction Mechanism ==

The mechanism of catalysis has not been established but must be similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes a phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref> The negatively charged oxygen of the last phosphate group on ADP nucleophilically attacks a phosphate of 1,3-phosphoglycerate. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form <scene name='Shane_Harmon_Sandbox/Atp/3'>hydrogen bonds</scene> with the enzyme. The 3 hydrogen bonds of ATP is favored over the 2 hydrogen bonds of ADP.

== Regulation ==

The role of PGK in glycolysis is very important for the production of ATP through substrate level phosphorylation. Regulation of this protein is thereby controlled by ATP or energy level of the cell. Recent research in frogs which can withstand freezing temperatures indicates that PGK is up regulated by the cold and more specifically by low levels of oxygen <ref> Shaobo, Wu et al. 2009. PGK1 expression responds to freezing, anoxia, and dehydration stresses in freeze tolerant wood frog, Rana sylvatica. Journal of Experimental Zoology. 311, 57-67 </ref> This makes logical sense as when oxygen is low, oxidative phosphorylation cannot occur. Thus without oxygen and oxidative phosphorylation, ATP levels begin to drop. In response to decreased ATP, PGK is up regulated to increase the amount of ATP produced by substrate level phosphorylation. It could therefore also be expected that ADP might act to inhibit or down regulated PGK expression.

Recent study of PGK has revolved around its function in tumor formation and growth. It has been shown that in addition to catalyzing its normal reaction of 1,3-Biphosphoglycerate and ADP to ATP and 3-Phosphoglycerate, PGK can also function to cleave disulfide bonds. Specifically, the review of sulfide bond cleavage indicates PGK has been shown to cleave disulfide bonds in the protein zymogen plasmin to produce the active form of the protein. The active form of plasmin is responsible for angiogenesis or blood vessel formation in tumors <ref> Hogg, PJ. 2002. Biological Regulation through protein disulfide bond cleavage. Redox Report. 7(2), 71-77. </ref> Without the formation of blood vessels to tumors nutrients are limited and tumor growth is limited. Once blood vessels are established growth can rapidly increase. The fact that tumor cells secrete PGK to allow blood vessel formation through the activation of the zymogen plasmin has important implications for understanding its regulation. If the regulation of PGK in tumor cells can be understood, it might be possible to inhibit the overproduction and secretion of PGK to limit angiogenesis in tumors.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-25T14:09:04Z

Shane Harmon: /* Mechanism of Phosphoglycerate Kinase(PGK) Catalysis */

=Mechanism of Phosphoglycerate Kinase(PGK) Catalysis=

== PGK in the Glycolysis Cycle ==

Phosphoglycerate kinase is a crucial enzyme in the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphoglycerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/2'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/4'>ATP</scene>. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously. {{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

== Structure ==

The overall structure of Phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of XXXXX amino acids. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP clssification of PGK is alpha and beta, indicating that its secondary strucutre is composed of roughly equal numbers alpha and beta sheets.

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrate molecules at the active sites, the proteins conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> Recent research indicates that the mechanism for closure of the two domains is a series of hydrogen bond interactions that occur upon binding of the substrates on both domains <ref>Vas, M, Varga, A et al. 2010. Insight into the Mechanism of of Domain Movements and their Role in Enzyme Function: Example of 3-Phosphoglycerate kinase. Current Protein and Peptide Science. Jan 21, 2010. (Epub ahead of publication).</ref>

== Kinetics ==

Given that Phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-Hofstee plots curve upward. One possible explanation for the non-linearity, negative cooperativity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 </ref>

== Reaction Mechanism ==

The mechanism of catalysis has not been established but must be similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes a phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref> The negatively charged oxygen of the last phosphate group on ADP nucleophilically attacks a phosphate of 1,3-phosphoglycerate. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form <scene name='Shane_Harmon_Sandbox/Atp/3'>hydrogen bonds</scene> with the enzyme. The 3 hydrogen bonds of ATP is favored over the 2 hydrogen bonds of ADP.

== Regulation ==

The role of PGK in glycolysis is very important for the production of ATP through substrate level phosphorylation. Regulation of this protein is thereby controlled by ATP or energy level of the cell. Recent research in frogs which can withstand freezing temperatures indicates that PGK is up regulated by the cold and more specifically by low levels of oxygen <ref> Shaobo, Wu et al. 2009. PGK1 expression responds to freezing, anoxia, and dehydration stresses in freeze tolerant wood frog, Rana sylvatica. Journal of Experimental Zoology. 311, 57-67 </ref> This makes logical sense as when oxygen is low, oxidative phosphorylation cannot occur. Thus without oxygen and oxidative phosphorylation, ATP levels begin to drop. In response to decreased ATP, PGK is up regulated to increase the amount of ATP produced by substrate level phosphorylation. It could therefore also be expected that ADP might act to inhibit or down regulated PGK expression.

Recent study of PGK has revolved around its function in tumor formation and growth. It has been shown that in addition to catalyzing its normal reaction of 1,3-Biphosphoglycerate and ADP to ATP and 3-Phosphoglycerate, PGK can also function to cleave disulfide bonds. Specifically, the review of sulfide bond cleavage indicates PGK has been shown to cleave disulfide bonds in the protein zymogen plasmin to produce the active form of the protein. The active form of plasmin is responsible for angiogenesis or blood vessel formation in tumors <ref> Hogg, PJ. 2002. Biological Regulation through protein disulfide bond cleavage. Redox Report. 7(2), 71-77. </ref> Without the formation of blood vessels to tumors nutrients are limited and tumor growth is limited. Once blood vessels are established growth can rapidly increase. The fact that tumor cells secrete PGK to allow blood vessel formation through the activation of the zymogen plasmin has important implications for understanding its regulation. If the regulation of PGK in tumor cells can be understood, it might be possible to inhibit the overproduction and secretion of PGK to limit angiogenesis in tumors.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-25T14:08:11Z

Shane Harmon: /* Mechanism of Phosphoglycerate Kinase(PGK) Catalysis */

Phosphoglycerate Kinase

2010-03-23T15:46:41Z

Shane Harmon: /* Mechanism of Phosphoglycerate Kinase(PGK) Catalysis */

=Mechanism of Phosphoglycerate Kinase(PGK) Catalysis=
Phosphoglycerate kinase is a crucial enzyme in the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphogylcerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/2'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/4'>ATP</scene>. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously.{{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

The overall structure of phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of XXXXX amino acids. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP clssification of PGK is alpha and beta, indicating that its secondary strucutre is composed of roughly equal numbers alpha and beta sheets.

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrate molecules at the active sites, the proteins conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> Rescent research indicates that the mechanism for closure of the two domains is a series of hydrogen bond interactions that occur upon binding of the substrates on both domains <ref>Vas, M, Varga, A et al. 2010. Insight into the Mechanism of of Domain Movements and their Role in Enzyme Function: Example of 3-Phosphoglycerate kinase. Current Protein and Peptide Science. Jan 21, 2010. (Epub ahead of publication).</ref>

Given that phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-hofstee plots curve upward. One possible explanation for the non-linearity, negative co-opertivity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 </ref>

The mechanism of catalysis has not been established but must be similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes a phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref> The negatively charged oxygen of the last phosphate group on ADP nucelophillically attacks a phosphate of 1,3-phosphoglycerate. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form <scene name='Shane_Harmon_Sandbox/Atp/3'>hydrogen bonds</scene> with the enzyme. The 3 hydrogen bonds of ATP is favored over the 2 hydrogen bonds of ADP.

The role of PGK in glycolysis is very important for the production of ATP through substrate level phosphorylation. Regulation of this protein is thereby controled by ATP or energy level of the cell. Recent research in frogs which can withstand freezing temperatures indicates that PGK is upregulated by the cold and more specifically by low levels of oxygen <ref> Shaobo, Wu et al. 2009. PGK1 expression responds to freezing, anoxia, and dehydration stresses in freeze tolerant wood frog, Rana sylvatica. Journal of Experimental Zoology. 311, 57-67 </ref> This makes logical sense as when oxygen is low, oxidative phosphorylation cannot occur. Thus without oxygen and oxidative phosphorylation, ATP levels begin to drop. In response to decreased ATP, PGK is upregulated to increase the amount of ATP produced by substrate level phosphorylation. It could therefore also be expected that ADP might act to inhibit or downregulated PGK expression.

Recent study of PGK has revolved around its function in tumor formation and growth. It has been shown that in addition to catalyzing its normal reaction of 1,3-Biphosphoglycerate and ADP to ATP and 3-Phosphoglycerate, PGK can also function to cleave disulfide bonds. Specifically, the review of sulfide bond cleavage indicates PGK has been shown to cleave disulfide bonds in the protein zymogen plasmin to produce the active form of the protein. The active form of plasmin is responsible for angiogenesis or blood vessel formation in tumors <ref> Hogg, PJ. 2002. Biological Regulation through protein disulfide bond cleavage. Redox Report. 7(2), 71-77. </ref> Without the formation of blood vessels to tumors nutrients are limited and tumor growth is limited. Once blood vessels are established growth can rapidly increase. The fact that tumor cells secrete PGK to allow blood vessel formation through the activation of the zymogen plasmin has important implications for understanding its regulation. If the regulation of PGK in tumor cells can understood, it might be possible to inhibit the overproduction and secretion of PGK to limit angiogenesis in tumors.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-23T15:22:57Z

Shane Harmon: /* Mechanism of Phosphoglycerate Kinase(PGK) Catalysis */

=Mechanism of Phosphoglycerate Kinase(PGK) Catalysis=
Phosphoglycerate kinase is a crucial enzyme in the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphogylcerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/1'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/1'>ATP</scene>. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously.{{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

The overall structure of phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of XXXXX amino acids. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP clssification of PGK is alpha and beta, indicating that its secondary strucutre is composed of roughly equal numbers alpha and beta sheets.

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrate molecules at the active sites, the proteins conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> Rescent research indicates that the mechanism for closure of the two domains is a series of hydrogen bond interactions that occur upon binding of the substrates on both domains <ref>Vas, M, Varga, A et al. 2010. Insight into the Mechanism of of Domain Movements and their Role in Enzyme Function: Example of 3-Phosphoglycerate kinase. Current Protein and Peptide Science. Jan 21, 2010. (Epub ahead of publication).</ref>

Given that phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-hofstee plots curve upward. One possible explanation for the non-linearity, negative co-opertivity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 </ref>

The mechanism of catalysis has not been established but must be similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes a phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref> The negatively charged oxygen of the last phosphate group on ADP nucelophillically attacks a phosphate of 1,3-phosphoglycerate. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form hydrogen bonds with the enzyme. The 3 hydrogen bonds of ATP is favored over the 2 hydrogen bonds of ADP.

The role of PGK in glycolysis is very important for the production of ATP through substrate level phosphorylation. Regulation of this protein is thereby controled by ATP or energy level of the cell. Recent research in frogs which can withstand freezing temperatures indicates that PGK is upregulated by the cold and more specifically by low levels of oxygen <ref> Shaobo, Wu et al. 2009. PGK1 expression responds to freezing, anoxia, and dehydration stresses in freeze tolerant wood frog, Rana sylvatica. Journal of Experimental Zoology. 311, 57-67 </ref> This makes logical sense as when oxygen is low, oxidative phosphorylation cannot occur. Thus without oxygen and oxidative phosphorylation, ATP levels begin to drop. In response to decreased ATP, PGK is upregulated to increase the amount of ATP produced by substrate level phosphorylation. It could therefore also be expected that ADP might act to inhibit or downregulated PGK expression.

Recent study of PGK has revolved around its function in tumor formation and growth. It has been shown that in addition to catalyzing its normal reaction of 1,3-Biphosphoglycerate and ADP to ATP and 3-Phosphoglycerate, PGK can also function to cleave disulfide bonds. Specifically, the review of sulfide bond cleavage indicates PGK has been shown to cleave disulfide bonds in the protein zymogen plasmin to produce the active form of the protein. The active form of plasmin is responsible for angiogenesis or blood vessel formation in tumors <ref> Hogg, PJ. 2002. Biological Regulation through protein disulfide bond cleavage. Redox Report. 7(2), 71-77. </ref> Without the formation of blood vessels to tumors nutrients are limited and tumor growth is limited. Once blood vessels are established growth can rapidly increase. The fact that tumor cells secrete PGK to allow blood vessel formation through the activation of the zymogen plasmin has important implications for understanding its regulation. If the regulation of PGK in tumor cells can understood, it might be possible to inhibit the overproduction and secretion of PGK to limit angiogenesis in tumors.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-23T15:17:19Z

Shane Harmon: /* Mechanism of Phosphoglycerate Kinase(PGK) Catalysis */

=Mechanism of Phosphoglycerate Kinase(PGK) Catalysis=
Phosphoglycerate kinase is a crucial enzyme in the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphogylcerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/1'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/1'>ATP</scene>. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously.{{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

The overall structure of phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of XXXXX amino acids. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP clssification of PGK is alpha and beta, indicating that its secondary strucutre is composed of roughly equal numbers alpha and beta sheets.

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrate molecules at the active sites, the proteins conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> Rescent research indicates that the mechanism for closure of the two domains is a series of hydrogen bond interactions that occur upon binding of the substrates on both domains <ref>Vas, M, Varga, A et al. 2010. Insight into the Mechanism of of Domain Movements and their Role in Enzyme Function: Example of 3-Phosphoglycerate kinase. Current Protein and Peptide Science. Jan 21, 2010. (Epub ahead of publication).</ref>

Given that phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-hofstee plots curve upward. One possible explanation for the non-linearity, negative co-opertivity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 </ref>

The mechanism of catalysis has not been established but must be similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes a phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref> The negatively charged oxygen of the last phosphate group on ADP nucelophillically attacks a phosphate of 1,3-phosphoglycerate. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form hydrogen bonds with the enzyme. The 3 hydrogen bonds of ATP is favored over the 2 hydrogen bonds of ADP.

The role of PGK in glycolysis is very important for the production of ATP through substrate level phosphorylation. Regulation of this protein is thereby controled by ATP or energy level of the cell. Recent research in frogs which can withstand freezing temperatures indicates that PGK is upregulated by the cold and more specifically by low levels of oxygen <ref> Shaobo, Wu et al. 2009. PGK1 expression responds to freezing, anoxia, and dehydration stresses in freeze tolerant wood frog, Rana sylvatica. Journal of Experimental Zoology. 311, 57-67 </ref> This makes logical sense as when oxygen is low, oxidative phosphorylation cannot occur. Thus without oxygen and oxidative phosphorylation, ATP levels begin to drop. In response to decreased ATP, PGK is upregulated to increase the amount of ATP produced by substrate level phosphorylation. It could therefore also be expected that ADP might act to inhibit or downregulated PGK expression.

Recent study of PGK has revolved around its function in tumor formation and growth. It has been shown that in addition to catalyzing its normal reaction of 1,3-Biphosphoglycerate and ADP to ATP and 3-Phosphoglycerate, PGK can also function to cleave disulfide bonds. Specifically, the review of sulfide bond cleavage indicates PGK has been shown to cleave disulfide bonds in the protein zymogen plasmin to produce the active form of the protein. The active form of plasmin is responsible for angiogenesis or blood vessel formation in tumors. Without the formation of blood vessels to tumors nutrients are limited and tumor growth is limited. Once blood vessels are established growth can rapidly increase.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-23T15:05:08Z

Shane Harmon: /* Mechanism of Phosphoglycerate Kinase(PGK) Catalysis */

=Mechanism of Phosphoglycerate Kinase(PGK) Catalysis=
Phosphoglycerate kinase is a crucial enzyme in the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphogylcerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/1'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/1'>ATP</scene>. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously.{{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

The overall structure of phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of XXXXX amino acids. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP clssification of PGK is alpha and beta, indicating that its secondary strucutre is composed of roughly equal numbers alpha and beta sheets.

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrate molecules at the active sites, the proteins conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> Rescent research indicates that the mechanism for closure of the two domains is a series of hydrogen bond interactions that occur upon binding of the substrates on both domains <ref>Vas, M, Varga, A et al. 2010. Insight into the Mechanism of of Domain Movements and their Role in Enzyme Function: Example of 3-Phosphoglycerate kinase. Current Protein and Peptide Science. Jan 21, 2010. (Epub ahead of publication).</ref>

Given that phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-hofstee plots curve upward. One possible explanation for the non-linearity, negative co-opertivity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 </ref>

The mechanism of catalysis has not been established but must be similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes a phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref> The negatively charged oxygen of the last phosphate group on ADP nucelophillically attacks a phosphate of 1,3-phosphoglycerate. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form hydrogen bonds with the enzyme. The 3 hydrogen bonds of ATP is favored over the 2 hydrogen bonds of ADP.

The role of PGK in glycolysis is very important for the production of ATP through substrate level phosphorylation. Regulation of this protein is thus a result of its function. Recent research in frogs which can withstand freezing temperatures indicates that PGK is upregulated by the cold and more specifically by low levels of oxygen <ref> Shaobo, Wu et al. 2009. PGK1 expression responds to freezing, anoxia, and dehydration stresses in freeze tolerant wood frog, Rana sylvatica. Journal of Experimental Zoology. 311, 57-67 </ref> This makes logical sense as when oxygen is low, oxidative phosphorylation cannot occur. Thus without oxygen and oxidative phosphorylation, ATP levels begin to drop. In response to decreased ATP, PGK is upregulated to increase the amount of ATP produced by substrate level phosphorylation. It could therefore be expected that ADP might act to inhibit or downregulated PGK expression.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-23T14:59:40Z

Shane Harmon: /* Mechanism of Phosphoglycerate Kinase(PGK) Catalysis */

=Mechanism of Phosphoglycerate Kinase(PGK) Catalysis=
Phosphoglycerate kinase is a crucial enzyme in the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphogylcerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/1'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/1'>ATP</scene>. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously.{{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

The overall structure of phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of XXXXX amino acids. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP clssification of PGK is alpha and beta, indicating that its secondary strucutre is composed of roughly equal numbers alpha and beta sheets.

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrate molecules at the active sites, the proteins conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> Rescent research indicates that the mechanism for closure of the two domains is a series of hydrogen bond interactions that occur upon binding of the substrates on both domains <ref>Vas, M, Varga, A et al. 2010. Insight into the Mechanism of of Domain Movements and their Role in Enzyme Function: Example of 3-Phosphoglycerate kinase. Current Protein and Peptide Science. Jan 21, 2010. (Epub ahead of publication).<ref>

Given that phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-hofstee plots curve upward. One possible explanation for the non-linearity, negative co-opertivity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 <ref>

The mechanism of catalysis has not been established but must be similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes a phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref> The negatively charged oxygen of the last phosphate group on ADP nucelophillically attacks a phosphate of 1,3-phosphoglycerate. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form hydrogen bonds with the enzyme. The 3 hydrogen bonds of ATP is favored over the 2 hydrogen bonds of ADP.

The role of PGK in glycolysis is very important for the production of ATP through substrate level phosphorylation. Regulation of this protein is thus a result of its function. Recent research in frogs which can withstand freezing temperatures indicates that PGK is upregulated by the cold and more specifically by low levels of oxygen <ref> Shaobo, Wu et al. 2009. PGK1 expression responds to freezing, anoxia, and dehydration stresses in freeze tolerant wood frog, Rana sylvatica. Journal of Experimental Zoology. 311, 57-67 <ref> This makes logical sense as when oxygen is low, oxidative phosphorylation cannot occur. Thus without oxygen and oxidative phosphorylation, ATP levels begin to drop. In response to decreased ATP, PGK is upregulated to increase the amount of ATP produced by substrate level phosphorylation. It could therefore be expected that ADP might act to inhibit or downregulated PGK expression.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-23T14:51:10Z

Shane Harmon: /* Mechanism of Phosphoglycerate Kinase(PGK) Catalysis */

=Mechanism of Phosphoglycerate Kinase(PGK) Catalysis=
Phosphogylcerate kinase is a crucial enzyme of the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphogylcerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/1'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/1'>ATP</scene>. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously.{{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

The overall structure of phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of XXXXX amino acids. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP clssification of PGK is alpha and beta, indicating that its secondary strucutre is composed of roughly equal numbers alpha and beta sheets.

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrate molecules at the active sites, the proteins conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref>

Given that phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-hofstee plots curve upward. One possible explanation for the non-linearity, negative co-opertivity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 <ref>

The mechanism of catalysis has not been established but must be similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes a phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref> The negatively charged oxygen of the last phosphate group on ADP nucelophillically attacks a phosphate of 1,3-phosphoglycerate. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form hydrogen bonds with the enzyme. The 3 hydrogen bonds of ATP is favored over the 2 hydrogen bonds of ADP.

The role of PGK in glycolysis is very important for the production of ATP through substrate level phosphorylation. Regulation of this protein is thus a result of its function. Recent research in frogs which can withstand freezing temperatures indicates that PGK is upregulated by the cold and more specifically by low levels of oxygen <ref> Shaobo, Wu et al. 2009. PGK1 expression responds to freezing, anoxia, and dehydration stresses in freeze tolerant wood frog, Rana sylvatica. Journal of Experimental Zoology. 311, 57-67 <ref> This makes logical sense as when oxygen is low, oxidative phosphorylation cannot occur. Thus without oxygen and oxidative phosphorylation, ATP levels begin to drop. In response to decreased ATP, PGK is upregulated to increase the amount of ATP produced by substrate level phosphorylation. It could therefore be expected that ADP might act to inhibit or downregulated PGK expression.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-23T14:35:24Z

Shane Harmon: /* Mechanism of Phosphoglycerate Kinase(PGK) Catalysis */

Phosphoglycerate Kinase

2010-03-23T14:26:44Z

Shane Harmon: /* Mechanism of Phosphoglycerate Kinase(PGK) Catalysis */

=Mechanism of Phosphoglycerate Kinase(PGK) Catalysis=
Phosphogylcerate kinase is a crucial enzyme of the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphogylcerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/1'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/1'>ATP</scene>. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously.{{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

The overall structure of phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of XXXXX amino acids. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP clssification of PGK is alpha and beta, indicating that its secondary strucutre is composed of roughly equal numbers alpha and beta sheets.

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrate molecules at the active sites, the proteins conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref>

Given that phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-hofstee plots curve upward. One possible explanation for the non-linearity, negative co-opertivity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516 <ref>

The mechanism of catalysis has not been established but must be similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes a phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed. <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref> The negatively charged oxygen of the last phosphate group on ADP nucelophillically attacks a phosphate of 1,3-phosphoglycerate. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form hydrogen bonds with the enzyme. The 3 hydrogen bonds of ATP is favored over the 2 hydrogen bonds of ADP.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-23T14:20:46Z

Shane Harmon: /* Mechanism of Phosphoglycerate Kinase(PGK) Catalysis */

=Mechanism of Phosphoglycerate Kinase(PGK) Catalysis=
Phosphogylcerate kinase is a crucial enzyme of the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphogylcerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/1'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/1'>ATP</scene>. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously.{{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

The overall structure of phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of XXXXX amino acids. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP clssification of PGK is alpha and beta, indicating that its secondary strucutre is composed of roughly equal numbers alpha and beta sheets.

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrate molecules at the active sites, the proteins conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref>

Given that phosphoglycerate kinase is a monomeric protein standard Michealis-Menton kinetics would be expected; however, this is not the case. Multiple experiments have shown that the data, when transformed into either double-reciprocal or Eadie-Hofstee plots is non-linear; Eadie-hofstee plots curve upward. One possible explanation for the non-linearity, negative co-opertivity, is ruled out because PGK does not have multiple subunits. In one study that conducted kinetic tests with a 1000 fold range of substrates, at the highest concentrations of substrate the rate was still increasing; this puts the Km value in the 2-5mM range <ref> Scopes, Robert. 1977. The Steady State Kinetics of Yeast Phosphoglycerate Kinase. European Journal of Biochemistry. 85, 503-516. <ref>

The mechanism of catalysis has not been established but must be similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes a phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed. <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref> The negatively charged oxygen of the last phosphate group on ADP nucelophillically attacks a phosphate of 1,3-phosphoglycerate. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form hydrogen bonds with the enzyme. The 3 hydrogen bonds of ATP is favored over the 2 hydrogen bonds of ADP.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-23T13:47:33Z

Shane Harmon: /* Mechanism of Phosphoglycerate Kinase(PGK) Catalysis */

Phosphoglycerate Kinase

2010-03-22T17:27:41Z

Shane Harmon:

=Mechanism of Phosphoglycerate Kinase(PGK) Catalysis=
Phosphogylcerate kinase is a crucial enzyme of the glycolysis cycle. This cycle is a series of ten reactions which ultimately breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphogylcerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/1'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/1'>ATP</scene>. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the less thermodynamically favored GADPH reaction of the cycle so both reactions occur spontaneously.{{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

The overall structure of phosphoglycerate kinase is very distinctive. It is a monomeric protein consisting of XXXXX amino acids. The structure is distinctly bilobed with a depressed region between the two lobes or domains. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene>. The SCOP clssification of PGK is alpha and beta, indicating that its secondary strucutre is composed of roughly equal numbers alpha and beta sheets.

The bilobed structure of PGK is very crucial in its catalytic function. The active site is broken into two pieces, one on the interior of each lobe or domain. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrate molecules at the active sites, the proteins conformation changes such that the two lobes of teh protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> When the two domains swing shut, a hydrophobic chamber free from water is established where the reaction can take place. The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref>

The SCOP classification of PGK is alpha and beta, indicating that is composed of roughly equal alpha and beta sheets. The tertiary stucture, or the overal structure, is that of a bilobed complex. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene> This enzyme has only one chain, thus its quaternary structure is that of a monomer.

The bilobed nature of the protein is very crucial in the its catlytic function. The active site is broken into two pieces, one on each interior lobe. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrates at the active sites, the protein's conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> This swinging shut of the protein creates an interior hydrophobic chamber that is free of water for the reaction to take place in.

The mechanism of catalysis has not been established but must be similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes a phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed. <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref> The negatively charged oxygen of the last phosphate group on ADP nucelophillically attacks a phosphate of 1,3-phosphoglycerate. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form hydrogen bonds with the enzyme. The 3 hydrogen bonds of ATP is favored over the 2 hydrogen bonds of ADP.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-22T17:15:10Z

Shane Harmon:

Phosphoglycerate Kinase

2010-03-22T17:13:30Z

Shane Harmon:

Phosphoglycerate Kinase

2010-03-01T05:34:50Z

Shane Harmon: /* Mechanism of Phosphoglycerate Kinase(PGK) Catalysis */

=Mechanism of Phosphoglycerate Kinase(PGK) Catalysis=
Phosphogylcerate kinase is a crucial enzyme of the glycolysis cycle. This cycle breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphogylcerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/1'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/1'>ATP</scene>. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the GADPH reaction of the cycle.{{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

The SCOP classification of PGK is alpha and beta, indicating that is composed of roughly equal alpha and beta sheets. The tertiary stucture, or the overal structure, is that of a bilobed complex. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene> This enzyme has only one chain, thus its quaternary structure is that of a monomer.

The bilobed nature of the protein is very crucial in the its catlytic function. The active site is broken into two pieces, one on each interior lobe. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrates at the active sites, the protein's conformation changes such that the two lobes of the protein swing together <ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> This swinging shut of the protein creates an interior hydrophobic chamber that is free of water for the reaction to take place in.

The mechanism of catalysis has not been established but must be similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes a phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed. <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref> The negatively charged oxygen of the last phosphate group on ADP nucelophillically attacks a phosphate of 1,3-phosphoglycerate. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form hydrogen bonds with the enzyme. The 3 hydrogen bonds of ATP is favored over the 2 hydrogen bonds of ADP.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-01T05:31:11Z

Shane Harmon: /* Mechanism of Phosphoglycerate Kinase(PGK) Catalysis */

Phosphoglycerate Kinase

2010-03-01T05:30:06Z

Shane Harmon:

=Mechanism of Phosphoglycerate Kinase(PGK) Catalysis=
Phosphogylcerate kinase is a crucial enzyme of the glycolysis cycle. This cycle breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphogylcerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/1'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/1'>ATP</scene>. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the GADPH reaction of the cycle.{{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

The SCOP classification of PGK is alpha and beta, indicating that is composed of roughly equal alpha and beta sheets. The tertiary stucture, or the overal structure, is that of a bilobed complex. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene> This enzyme has only one chain, thus its quaternary structure is that of a monomer.

The bilobed nature of the protein is very crucial in the its catlytic function. The active site is broken into two pieces, one on each interior lobe. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrates at the active sites, the protein's conformation changes such that the two lobes of the protein swing together (<ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref> This swinging shut of the protein creates an interior hydrophobic chamber that is free of water for the reaction to take place in.

The mechanism of catalysis has not been established but must be similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes a phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed. <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref> The negatively charged oxygen of the last phosphate group on ADP nucelophillically attacks a phosphate of 1,3-phosphoglycerate. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form hydrogen bonds with the enzyme. The 3 hydrogen bonds of ATP is favored over the 2 hydrogen bonds of ADP.

Replace the PDB id (use lowercase!) after the STRUCTURE_ and after PDB= to load
and display another structure.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-01T05:28:09Z

Shane Harmon:

=Mechanism of Phosphoglycerate Kinase(PGK) Catalysis=
Phosphogylcerate kinase is a crucial enzyme of the glycolysis cycle. This cycle breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphogylcerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/1'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/1'>ATP</scene>. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the GADPH reaction of the cycle.{{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

The SCOP classification of PGK is alpha and beta, indicating that is composed of roughly equal alpha and beta sheets. The tertiary stucture, or the overal structure, is that of a bilobed complex. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene> This enzyme has only one chain, thus its quaternary structure is that of a monomer.

The bilobed nature of the protein is very crucial in the its catlytic function. The active site is broken into two pieces, one on each interior lobe. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrates at the active sites, the protein's conformation changes such that the two lobes of the protein swing together (<ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref>== <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> This swinging shut of the protein creates an interior hydrophobic chamber that is free of water for the reaction to take place in.

The mechanism of catalysis has not been established but must be similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes a phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed. <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref> The negatively charged oxygen of the last phosphate group on ADP nucelophillically attacks a phosphate of 1,3-phosphoglycerate. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form hydrogen bonds with the enzyme. The 3 hydrogen bonds of ATP is favored over the 2 hydrogen bonds of ADP.

Replace the PDB id (use lowercase!) after the STRUCTURE_ and after PDB= to load
and display another structure.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-01T05:27:24Z

Shane Harmon:

Mechanism of Phosphoglycerate Kinase(PGK) Catalysis==
Phosphogylcerate kinase is a crucial enzyme of the glycolysis cycle. This cycle breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphogylcerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/1'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/1'>ATP</scene>. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the GADPH reaction of the cycle.{{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

The SCOP classification of PGK is alpha and beta, indicating that is composed of roughly equal alpha and beta sheets. The tertiary stucture, or the overal structure, is that of a bilobed complex. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene> This enzyme has only one chain, thus its quaternary structure is that of a monomer.

The bilobed nature of the protein is very crucial in the its catlytic function. The active site is broken into two pieces, one on each interior lobe. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrates at the active sites, the protein's conformation changes such that the two lobes of the protein swing together (<ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref>== <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> This swinging shut of the protein creates an interior hydrophobic chamber that is free of water for the reaction to take place in.

The mechanism of catalysis has not been established but must be similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes a phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed. <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref> The negatively charged oxygen of the last phosphate group on ADP nucelophillically attacks a phosphate of 1,3-phosphoglycerate. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form hydrogen bonds with the enzyme. The 3 hydrogen bonds of ATP is favored over the 2 hydrogen bonds of ADP.

Replace the PDB id (use lowercase!) after the STRUCTURE_ and after PDB= to load
and display another structure.

<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-01T05:16:30Z

Shane Harmon: /* Mechanism of Phosphoglycerate Kinase(PGK) Catalysis */

<ref>Blake and Rice. 1981. Phosphoglycerate kinase. Philosophical Transactions of the Royal Society of London. 293:93-104.</ref>==Mechanism of Phosphoglycerate Kinase(PGK) Catalysis==
Phosphogylcerate kinase is a crucial enzyme of the glycolysis cycle. This cycle breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphogylcerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/1'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/1'>ATP</scene>. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the GADPH reaction of the cycle.{{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

The SCOP classification of PGK is alpha and beta, indicating that is composed of roughly equal alpha and beta sheets. The tertiary stucture, or the overal structure, is that of a bilobed complex. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene> This enzyme has only one chain, thus its quaternary structure is that of a monomer.

The bilobed nature of the protein is very crucial in the its catlytic function. The active site is broken into two pieces, one on each interior lobe. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrates at the active sites, the protein's conformation changes such that the two lobes of the protein swing together (<ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> This swinging shut of the protein creates an interior hydrophobic chamber that is free of water for the reaction to take place in.

The mechanism of catalysis has not been established but must be similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes a phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed. <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref> The negatively charged oxygen of the last phosphate group on ADP nucelophillically attacks a phosphate of 1,3-phosphoglycerate. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form hydrogen bonds with the enzyme. The 3 hydrogen bonds of ATP is favored over the 2 hydrogen bonds of ADP.

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<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-01T05:11:37Z

Shane Harmon: /* Mechanism of Phosphoglycerate Kinase(PGK) Catalysis */

==Mechanism of Phosphoglycerate Kinase(PGK) Catalysis==
Phosphogylcerate kinase is a crucial enzyme of the glycolysis cycle. This cycle breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphogylcerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/1'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/1'>ATP</scene>. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the GADPH reaction of the cycle.{{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

The SCOP classification of PGK is alpha and beta, indicating that is composed of roughly equal alpha and beta sheets. The tertiary stucture, or the overal structure, is that of a bilobed complex. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene> This enzyme has only one chain, thus its quaternary structure is that of a monomer.

The bilobed nature of the protein is very crucial in the its catlytic function. The active site is broken into two pieces, one on each interior lobe. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrates at the active sites, the protein's conformation changes such that the two lobes of the protein swing together (<ref>Voet, Donald et al. 2008. Fundamentals of Biochemistry. 3rd ed. 499 </ref> The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> This swinging shut of the protein creates an interior hydrophobic chamber that is free of water for the reaction to take place in.

The mechanism of catalysis has not been established but must be similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes a phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed. <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref> The negatively charged oxygen of the last phosphate group on ADP nucelophillically attacks a phosphate of 1,3-phosphoglycerate. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form hydrogen bonds with the enzyme. The 3 hydrogen bonds of ATP is favored over the 2 hydrogen bonds of ADP.

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<references/>
<ref></ref>

Phosphoglycerate Kinase

2010-03-01T05:06:43Z

Shane Harmon: /* Mechanism of Phosphoglycerate Kinase(PGK) Catalysis */

==Mechanism of Phosphoglycerate Kinase(PGK) Catalysis==
Phosphogylcerate kinase is a crucial enzyme of the glycolysis cycle. This cycle breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphogylcerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/1'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/1'>ATP</scene>. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the GADPH reaction of the cycle.{{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

The SCOP classification of PGK is alpha and beta, indicating that is composed of roughly equal alpha and beta sheets. The tertiary stucture, or the overal structure, is that of a bilobed complex. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene> This enzyme has only one chain, thus its quaternary structure is that of a monomer.

The bilobed nature of the protein is very crucial in the its catlytic function. The active site is broken into two pieces, one on each interior lobe. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrates at the active sites, the protein's conformation changes such that the two lobes of the protein swing together. The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> This swinging shut of the protein creates an interior hydrophobic chamber that is free of water for the reaction to take place in.

The mechanism of catalysis has not been established but must be similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes a phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed. <ref>Harnan, G. et al. 1992. Domain Motions in Phosphoglycerate Kinase: Determination of Interdomain Distance Distribution by Site Specific Labeling and Time Resolved Flourescense Energy Transfer. PNAS. 89:11764-11768.</ref> The negatively charged oxygen of the last phosphate group on ADP nucelophillically attacks a phosphate of 1,3-phosphoglycerate. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form hydrogen bonds with the enzyme. The 3 hydrogen bonds of ATP is favored over the 2 hydrogen bonds of ADP.

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<references/>

Phosphoglycerate Kinase

2010-03-01T04:57:18Z

Shane Harmon: /* Mechanism of Phosphoglycerate Kinase(PGK) Catalysis */

==Mechanism of Phosphoglycerate Kinase(PGK) Catalysis==
Phosphogylcerate kinase is a crucial enzyme of the glycolysis cycle. This cycle breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphogylcerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/1'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/1'>ATP</scene>. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the GADPH reaction of the cycle.{{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

The SCOP classification of PGK is alpha and beta, indicating that is composed of roughly equal alpha and beta sheets. The tertiary stucture, or the overal structure, is that of a bilobed complex. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene> This enzyme has only one chain, thus its quaternary structure is that of a monomer.

The bilobed nature of the protein is very crucial in the its catlytic function. The active site is broken into two pieces, one on each interior lobe. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrates at the active sites, the protein's conformation changes such that the two lobes of the protein swing together. The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> This swinging shut of the protein creates an interior hydrophobic chamber that is free of water for the reaction to take place in.

The mechanism of catalysis has not been established but must be similar to that of hexokinase. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose and has a very similar structure and conformational change via a hinge. PGK has a similar function except it catalyzes the transfer of a phosphate to form ATP instead of using ATP. The reaction of PGK removes a phosphate group from the intermediate molecule, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed. The negatively charged oxygen of the last phosphate group on ADP nucelophillically attacks a phosphate of 1,3-phosphoglycerate. The product, ATP, is favored because it's negatively charged oxygens of the 3 phosphates form hydrogen bonds with the enzyme. The 3 hydrogen bonds of ATP is favored over the 2 hydrogen bonds of ADP.

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Phosphoglycerate Kinase

2010-03-01T04:50:31Z

Shane Harmon: /* Mechanism of Phosphoglycerate Kinase(PGK) Catalysis */

==Mechanism of Phosphoglycerate Kinase(PGK) Catalysis==
Phosphogylcerate kinase is a crucial enzyme of the glycolysis cycle. This cycle breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphogylcerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/1'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/1'>ATP</scene>. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, or an ATPase. The reaction is highly exergonic allowing it to be coupled with the GADPH reaction of the cycle.{{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

The SCOP classification of PGK is alpha and beta, indicating that is composed of roughly equal alpha and beta sheets. The tertiary stucture, or the overal structure, is that of a bilobed complex. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene> This enzyme has only one chain, thus its quaternary structure is that of a monomer.

The bilobed nature of the protein is very crucial in the its catlytic function. The active site is broken into two pieces, one on each interior lobe. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrates at the active sites, the protein's conformation changes such that the two lobes of the protein swing together. The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> This swinging shut of the protein creates an interior hydrophobic chamber that is free of water for the reaction to take place in.

The mechanism of catalysis must be similar to that of hexokinase since the two enzymes both have a similar function and structure. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose. The reaction of PGK removes a phosphate group from the intermediate molecue, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed. The negatively charged oxygen of the last phosphate group on ADP nucelophillically attacks a phosphate of 1,3-phosphoglycerate.

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Phosphoglycerate Kinase

2010-03-01T04:49:19Z

Shane Harmon: /* Mechanism of Phosphoglycerate Kinase(PGK) Catalysis */

==Mechanism of Phosphoglycerate Kinase(PGK) Catalysis==
Phosphogylcerate kinase is a crucial enzyme of the glycolysis cycle. This cycle breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphogylcerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce <scene name='Shane_Harmon_Sandbox/Product/1'>3-Phosphoglycerate</scene> and <scene name='Shane_Harmon_Sandbox/Atp/1'>ATP</scene>. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, and an ATPase. The reaction is highly exergonic allowing it to be coupled with the GADPH reaction of the cycle.{{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

The SCOP classification of PGK is alpha and beta, indicating that is composed of roughly equal alpha and beta sheets. The tertiary stucture, or the overal structure, is that of a bilobed complex. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene> This enzyme has only one chain, thus its quaternary structure is that of a monomer.

The bilobed nature of the protein is very crucial in the its catlytic function. The active site is broken into two pieces, one on each interior lobe. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrates at the active sites, the protein's conformation changes such that the two lobes of the protein swing together. The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> This swinging shut of the protein creates an interior hydrophobic chamber that is free of water for the reaction to take place in.

The mechanism of catalysis must be similar to that of hexokinase since the two enzymes both have a similar function and structure. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose. The reaction of PGK removes a phosphate group from the intermediate molecue, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed. The negatively charged oxygen of the last phosphate group on ADP nucelophillically attacks a phosphate of 1,3-phosphoglycerate.

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Phosphoglycerate Kinase

2010-03-01T04:40:06Z

Shane Harmon: /* Mechanism of Phosphoglycerate Kinase(PGK) Catalysis */

==Mechanism of Phosphoglycerate Kinase(PGK) Catalysis==
Phosphogylcerate kinase is a crucial enzyme of the glycolysis cycle. This cycle breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphogylcerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce 3-Phosphoglycerate and <scene name='Shane_Harmon_Sandbox/Atp/1'>Ligand</scene>. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, and an ATPase. The reaction is highly exergonic allowing it to be coupled with the GADPH reaction of the cycle.{{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

The SCOP classification of PGK is alpha and beta, indicating that is composed of roughly equal alpha and beta sheets. The tertiary stucture, or the overal structure, is that of a bilobed complex. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene> This enzyme has only one chain, thus its quaternary structure is that of a monomer.

The bilobed nature of the protein is very crucial in the its catlytic function. The active site is broken into two pieces, one on each interior lobe. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrates at the active sites, the protein's conformation changes such that the two lobes of the protein swing together. The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between <scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> This swinging shut of the protein creates an interior hydrophobic chamber that is free of water for the reaction to take place in.

The mechanism of catalysis must be similar to that of hexokinase since the two enzymes both have a similar function and structure. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose. The reaction of PGK removes a phosphate group from the intermediate molecue, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed. The negatively charged oxygen of the last phosphate group on ADP nucelophillically attacks a phosphate of 1,3-phosphoglycerate.

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Phosphoglycerate Kinase

2010-03-01T04:32:16Z

Shane Harmon: /* Mechanism of Phosphoglycerate Kinase(PGK) Catalysis */

==Mechanism of Phosphoglycerate Kinase(PGK) Catalysis==
Phosphogylcerate kinase is a crucial enzyme of the glycolysis cycle. This cycle breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphogylcerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce 3-Phosphoglycerate and ATP. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, and an ATPase. The reaction is highly exergonic allowing it to be coupled with the GADPH reaction of the cycle.{{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

The SCOP classification of PGK is alpha and beta, indicating that is composed of roughly equal alpha and beta sheets. The tertiary stucture, or the overal structure, is that of a bilobed complex. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene> This enzyme has only one chain, thus its quaternary structure is that of a monomer.

The bilobed nature of the protein is very crucial in the its catlytic function. The active site is broken into two pieces, one on each interior lobe. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrates at the active sites, the protein's conformation changes such that the two lobes of the protein swing together. The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between ARG62 and ASP200.<scene name='Shane_Harmon_Sandbox/Salt_bridge/1'>ARG62 and ASP200.</scene> This swinging shut of the protein creates an interior hydrophobic chamber that is free of water for the reaction to take place in.

<scene name='Shane_Harmon_Sandbox/Atp/1'>Ligand</scene>

The mechanism of catalysis must be similar to that of hexokinase since the two enzymes both have a similar function and structure. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose. The reaction of PGK removes a phosphate group from the intermediate molecue, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed. The negatively charged oxygen of the last phosphate group on ADP nucelophillically attacks a phosphate of 1,3-phosphoglycerate.

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Phosphoglycerate Kinase

2010-03-01T04:18:51Z

Shane Harmon: /* Mechanism of Phosphoglycerate Kinase(PGK) Catalysis */

==Mechanism of Phosphoglycerate Kinase(PGK) Catalysis==
Phosphogylcerate kinase is a crucial enzyme of the glycolysis cycle. This cycle breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphogylcerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce 3-Phosphoglycerate and ATP. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, and an ATPase. The reaction is highly exergonic allowing it to be coupled with the GADPH reaction of the cycle.{{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

The SCOP classification of PGK is alpha and beta, indicating that is composed of roughly equal alpha and beta sheets. The tertiary stucture, or the overal structure, is that of a bilobed complex. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene> This enzyme has only one chain, thus its quaternary structure is that of a monomer.

The bilobed nature of the protein is very crucial in the its catlytic function. The active site is broken into two pieces, one on each interior lobe. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrates at the active sites, the protein's conformation changes such that the two lobes of the protein swing together. The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between ARG62 and ASP200. This swinging shut of the protein creates an interior hydrophobic chamber that is free of water for the reaction to take place in.

<scene name='Shane_Harmon_Sandbox/Atp/1'>Ligand</scene>

The mechanism of catalysis must be similar to that of hexokinase since the two enzymes both have a similar function and structure. Hexokinase catalyzes the removal of a phosphate group from ATP to glucose. The reaction of PGK removes a phosphate group from the intermediate molecue, 1,3-biphosphoglycerate and transfers it to ADP to form ATP. Once the substrates bind to the active sites, the protein domains swing shut forcing the substrates into correct position for the reaction to proceed. The negatively charged oxygen of the last phosphate group on ADP nucelophillically attacks a phosphate of 1,3-phosphoglycerate.

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Phosphoglycerate Kinase

2010-03-01T04:08:48Z

Shane Harmon: /* This is a placeholder */

Phosphoglycerate Kinase

2010-03-01T03:39:48Z

Shane Harmon: /* This is a placeholder */

==This is a placeholder==
Phosphogylcerate kinase is a crucial enzyme of the glycolysis cycle. This cycle breaks down glucose into pyruvate while generating 2 NADH and 2 ATP molecules. Phosphogylcerate kinase is the seventh enzyme in the cycle which catalyzes the reaction of 1,3-Biphosphoglycerate and ADP to produce 3-Phosphoglycerate and ATP. This method for ATP production is known as substrate level phosphorylation because it produces energy storing ATP molecules with out the use of oxygen, NADH, and an ATPase. The reaction is highly exergonic allowing it to be coupled with the GADPH reaction of the cycle.{{STRUCTURE_3cin | PDB=3PGK | SCENE= }}

The SCOP classification of PGK is alpha and beta, indicating that is composed of roughly equal alpha and beta sheets. The tertiary stucture, or the overal structure, is that of a bilobed complex. The lobes/domains are clearly connected at only two locations:<scene name='Shane_Harmon_Sandbox/Domain_links/1'>Beta Sheet L, Residues 189-202 and between Alpha Helix 14 and 15, Residues 404-408</scene> This enzyme has only one chain, thus its quaternary structure is that of a monomer.

The bilobed nature of the protein is very crucial in the its catlytic function. The active site is broken into two pieces, one on each interior lobe. On one site the ADP-Mg2+ substrate binds and on the other lobe the 1,3-Biphosphoglycerate substrate binds. Upon binding of both substrates at the active sites, the protein's conformation changes such that the two lobes of the protein swing together. The hinge for this conformational change is beta sheet L and the new conformation is formed via a salt bridge between ARG62 and ASP200. This swinging shut of the protein creates an interior hydrophobic chamber that is free of water for the reaction to take place in.

<scene name='Shane_Harmon_Sandbox/Atp/1'>Ligand</scene>

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