61473-66-3Relevant articles and documents
Chemical Modification of Cystein Residues of L-Methionine γ-Lyase
Nakayama, Toru,Esaki, Nobuyoshi,Tanaka, Hidehiko,Soda, Kenji
, p. 177 - 184 (2007/10/02)
The L-methionine γ-lyase of Pseudomonas putida contains 16 cysteinyl residues per mol of enzyme, which is composed of four identical polypeptide chains.Eight of these residues are buried inside the enzyme structure; they reacted with 5,5'-dithiobis(2-nitrobenzoic acid) only after the enzyme was denatured.In the native enzyme, two of them were cyanylated with 2-nitro-5-thiocyanobenzoic acid with concomitant inactivation of the enzyme.Both the cyanylation and the inactivation proceeded through biphasic pseudo-first order kinetics.Cyanylation of about 2 mol of cystein residues per mol of enzyme practically abolished the α,γ-elimination activity: the enzyme shows a half-of-the-sites reactivity.L-Norleucine, a competetive inhibitor, which binds to the active site, strongly retarded the reaction of the essential cystein residue with 2-nitro-5-thiocyanobenzoic acid, and protected the enzyme from inactivation.The cyanylated enzyme was reactivated with 2-mercaptoethanol formed in situ from S-(β-hydroxyethyl)-L-homocystein by its remaining activity.Added 2-mercaptoethanol was much less effective than that formed in situ.These suggest that the essential cystein residue is at or near the active site.The cyanylated enzyme was substantially inactive (residual activity, 5percent) for the α,γ-elimination, but 40percent active for the α,β-elimination.Kinetic analyses revealed that the affinity of enzyme for the substrates, particularly for long straight-chain amino acids, was decreased greatly by cyanylation.The cyanylated cystein residue was found to be located at a position of approximately 20percent of the total lenght of the polypeptide from its NH2-terminus.
