Site‐directed mutagenesis of histidine 62 in the ‘basic patch’ region of yeast phosphoglycerate kinase

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Abstract
Site‐directed mutagenesis has been used to produce a mutant form of yeast phosphoglycerate kinase (PGK) in which the ‘basic patch’ residue His 62 has been replaced by a glutamine residue. Using 1H‐NMR spectroscopy, it was found that 3‐phosphoglycerate (3‐PG) binding to the mutant protein induces the same conformational effects as for wild‐type PGK, although the affinity was reduced by 2‐ to 3‐fold. Kinetic studies show both K m for 3‐PG and V max to be increased by ~ 2‐fold relative to the wild‐type enzyme. These data are consistent with the suggestion that His 62 assists in the binding of the substrate to the enzyme.