Christopher Vachon Sandbox: Difference between revisions

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It is important to note that the phosphate group that is placed on C2 is not the same phosphate group that was initially on C3.  
It is important to note that the phosphate group that is placed on C2 is not the same phosphate group that was initially on C3.  
In order to understand how PGAM catalyzes this reaction, an explanation of its active site is imperative.  The most important residues in this enzyme include <scene name='Christopher_Vachon_Sandbox/His_8_good/2'>His 8 and 181</scene> with imidazole groups which are in close proximity to carbons 2 and 3 in the substrate. Based on crystallography experiments, the active site where these histidine residues reside lies at the bottom of a deep groove in each subunit. <ref>S., Winn I., Fothergill A. L., Harkins N. R., and Watson C. H. "Structure and Activity of Phosphoglycerate Mutase." Sciences 293.1063 (1981): 121-30. Print.</ref>  The sites in each subunit, whether the enzyme is a homodimer or homotetramer, are well separated.  The active enzyme contains a phosphoryl group attached to His 8.  This phosphoryl group is what is transferred to C2 of the substrate, resulting in an intermediate 2,3-bisphosphoglycerate-enzyme complex.  Thus there is a <scene name='Christopher_Vachon_Sandbox/Good_active_site_scene/4'>covalently attached phosphate</scene> in the active monomer. <ref>Voet, Donald, Judith G. Voet, and Charlotte W. Pratt. Fundamentals of Biochemistry Life at the Molecular Level. New York: John Wiley & Sons, 2008. Print.</ref> The phosphate group on C3 of the substrate is then transferred back onto His 8, thus regenerating the active form of the enzyme.   
In order to understand how PGAM catalyzes this reaction, an explanation of its active site is imperative.  The most important residues in this enzyme include <scene name='Christopher_Vachon_Sandbox/His_8_good/2'>His 8 and 179</scene> with imidazole groups which are in close proximity to carbons 2 and 3 in the substrate.  His-8 is phosphorylated during during catalysis, and it is likely that His-179 acts as the proton donor/acceptor <ref>Rose, Z.B. (1980) Adv. Enzymol. Relat. Areas Mol. Biol. 51, 211-253</ref>. Based on crystallography experiments, the active site where these histidine residues reside lies at the bottom of a deep groove in each subunit. <ref>S., Winn I., Fothergill A. L., Harkins N. R., and Watson C. H. "Structure and Activity of Phosphoglycerate Mutase." Sciences 293.1063 (1981): 121-30. Print.</ref>  The sites in each subunit, whether the enzyme is a homodimer or homotetramer, are well separated.  The active enzyme contains a phosphoryl group attached to His 8.  This phosphoryl group is what is transferred to C2 of the substrate, resulting in an intermediate 2,3-bisphosphoglycerate-enzyme complex.  Thus there is a <scene name='Christopher_Vachon_Sandbox/Good_active_site_scene/4'>covalently attached phosphate</scene> in the active monomer. <ref>Voet, Donald, Judith G. Voet, and Charlotte W. Pratt. Fundamentals of Biochemistry Life at the Molecular Level. New York: John Wiley & Sons, 2008. Print.</ref> The phosphate group on C3 of the substrate is then transferred back onto His 8, thus regenerating the active form of the enzyme.  
   
In addition to the importance of the two histidine residues in the active site, the amino acids that line the <scene name='Christopher_Vachon_Sandbox/Good_active_site_scene/3'>active site</scene> are also functionally important.  These residues include H179, H8, E15, S11, T20, R59, and E86.<ref>Voet, Donald, Judith G. Voet, and Charlotte W. Pratt. Fundamentals of Biochemistry Life at the Molecular Level. New York: John Wiley & Sons, 2008. Print.</ref> Several positively charged residues line the active site pocket, which are usually <scene name='Christopher_Vachon_Sandbox/Arginine_residues/1'>arginine residues</scene>. <ref>S., Winn I., Fothergill A. L., Harkins N. R., and Watson C. H. "Structure and Activity of Phosphoglycerate Mutase." Sciences 293.1063 (1981): 121-30. Print.</ref>  This structure is logical for its function because the enzyme binds a negatively charged substrate, thus a positively charged groove fosters tight binding with a negative substrate.  The third and final important aspect of the active site is the presence of <scene name='Christopher_Vachon_Sandbox/Glutamate_residues/1'>glutamate residues 15 and 86</scene>.<ref>S., Winn I., Fothergill A. L., Harkins N. R., and Watson C. H. "Structure and Activity of Phosphoglycerate Mutase." Sciences 293.1063 (1981): 121-30. Print.</ref>  It is suggested that the carboxyl groups of these amino acid residues act as proton-withdrawing groups as they flank both sides of the substrate.
In addition to the importance of the two histidine residues in the active site, the amino acids that line the <scene name='Christopher_Vachon_Sandbox/Good_active_site_scene/3'>active site</scene> are also functionally important.  These residues include H179, H8, E15, S11, T20, R59, and E86.<ref>Voet, Donald, Judith G. Voet, and Charlotte W. Pratt. Fundamentals of Biochemistry Life at the Molecular Level. New York: John Wiley & Sons, 2008. Print.</ref> Several positively charged residues line the active site pocket, which are usually <scene name='Christopher_Vachon_Sandbox/Arginine_residues/1'>arginine residues</scene>. <ref>S., Winn I., Fothergill A. L., Harkins N. R., and Watson C. H. "Structure and Activity of Phosphoglycerate Mutase." Sciences 293.1063 (1981): 121-30. Print.</ref>  This structure is logical for its function because the enzyme binds a negatively charged substrate, thus a positively charged groove fosters tight binding with a negative substrate.  The third and final important aspect of the active site is the presence of <scene name='Christopher_Vachon_Sandbox/Glutamate_residues/1'>glutamate residues 15 and 86</scene>.<ref>S., Winn I., Fothergill A. L., Harkins N. R., and Watson C. H. "Structure and Activity of Phosphoglycerate Mutase." Sciences 293.1063 (1981): 121-30. Print.</ref>  It is suggested that the carboxyl groups of these amino acid residues act as proton-withdrawing groups as they flank both sides of the substrate.
In terms of regulation, competitive inhibitors resemble the negatively charged substrate and bind to the active site.  Such inhibitors include inositol hexakisphosphate and benzene hexacarboxylate.  <ref>Rigden, D. J.; Walter, R. A.; Phillips, S. E. V.; Fothergill-Gilmore, L. A.Polyanionic inhibitors of phosphoglycerate mutase: combined structural and biochemical analysis  J. Mol. Biol. 1999, 289, 691– 699</ref>
 
In terms of regulation, competitive inhibitors resemble the negatively charged substrate and bind to the active site.  Such inhibitors include inositol hexakisphosphate and benzene hexacarboxylate.  <ref>Rigden, D. J.; Walter, R. A.; Phillips, S. E. V.; Fothergill-Gilmore, L. A.Polyanionic inhibitors of phosphoglycerate mutase: combined structural and biochemical analysis  J. Mol. Biol. 1999, 289, 691– 699</ref>  Additionally, the phosphomethyl analogue of 3-phosphoglycerate (2-hydroxy-4-phosphonobutanoate) is a potent inhibitor of phosphoglycerate mutase. <ref>McAleese, S.M., Fothergill-Gilmore, L.A.&Dixon, H.B.F. (1985) Biochem. J. 230, 535-542 </ref>




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