PMID:10026144
| Citation |
Grant, GA, Kim, SJ, Xu, XL and Hu, Z (1999) The contribution of adjacent subunits to the active sites of D-3-phosphoglycerate dehydrogenase. J. Biol. Chem. 274:5357-61 |
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| Abstract |
D-3-Phosphoglycerate dehydrogenase (PGDH) from Escherichia coli is allosterically inhibited by L-serine, the end product of its metabolic pathway. Previous results have shown that inhibition by serine has a large effect on Vmax and only a small or negligible effect on Km. PGDH is thus classified as a V-type allosteric enzyme. In this study, the active site of PGDH has been studied by site-directed mutagenesis to assess the role of certain residues in substrate binding and catalysis. These consist of a group of cationic residues (Arg-240, Arg-60, Arg-62, Lys-39, and Lys-141') that potentially form an electrostatic environment for the binding of the negatively charged substrate, as well as the only tryptophan residue found in PGDH and which fits into a hydrophobic pocket immediately adjacent to the active site histidine residue. Interestingly, Trp-139' and Lys-141' are part of the polypeptide chain of the subunit that is adjacent to the active site. The results of mutating these residues show that Arg-240, Arg-60, Arg-62, and Lys-141' play distinct roles in the binding of the substrate to the active site. Mutants of Trp-139' show that this residue may play a role in stabilizing the catalytic center of the enzyme. Furthermore, these mutants appear to have a significant effect on the cooperativity of serine inhibition and suggest a possible role for Trp-139' in the cooperative interactions between subunits. |
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| Keywords |
Amino Acid Substitution; Binding Sites; Carbohydrate Dehydrogenases/chemistry; Carbohydrate Dehydrogenases/genetics; Carbohydrate Dehydrogenases/metabolism; Catalysis; Cations; Escherichia coli/enzymology; Kinetics; Models, Molecular; Mutagenesis, Site-Directed; Phosphoglycerate Dehydrogenase |
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