Role of Tyrosine 114 of L-Methionine γ-lyase from Pseudomonas putida

概要

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L-Methionine γ-lyase from Pseudomonas putida has a conserved tyrosine residue (Tyr114) in the active site as in all known sequences of γ-family pyridoxal 5'-phosphate dependent enzymes. A mutant form of L-methionine γ-lyase in which Tyr114 was replaced by phenylalanine (Y114F) resulted in 910-fold decrease in k_<cat> for α, γ-elimination of L-methionine, while the K_m remained the same as the wild type enzyme. The Y114F mutant had the reduced k_<cat> by only 28-and 16-fold for substrates with an electron-withdrawing group at the γ-position, namely O-acetyl-L-homoserine and L-methionine sulfone, respectively, and also the similar reduction of k_<cat> for α, β-elimination and deamination substrates. The hydrogen exchange reactions of substrate and the spectral changes of the substrate-enzyme complex catalyzed by the mutant enzyme suggested that γ-elimination process for L-methionine is the rate-limiting determination step in α, γ-elimination overall reaction of the Y114F mutant. These results indicate that Tyr114 of L-methionine γ-lyase is important in γ-elimination of the substrate.
社団法人日本農芸化学会の論文
2000-11-23