title: Arg305 of Streptomyces L-glutamate oxidase plays a crucial role for substrate recognition.
abstract: Recently, we have solved the crystal structure of L-glutamate oxidase (LGOX) from Streptomyces sp. X-119-6 (PDB code: 2E1M), the substrate specificity of which is strict toward L-glutamate. By a docking simulation using L-glutamate and structure of LGOX, we selected three residues, Arg305, His312, and Trp564 as candidates of the residues associating with recognition of L-glutamate. The activity of LGOX toward L-glutamate was significantly reduced by substitution of selected residues with Ala. However, the enzyme, Arg305 of which was substituted with Ala, exhibited catalytic activity toward various L-amino acids. To investigate the role of Arg305 in substrate specificity, we constructed Arg305 variants of LGOX. In all mutants, the substrate specificity of LGOX was markedly changed by the mutation. The results of kinetics and pH dependence on activity indicate that Arg305 of LGOX is associated with the interaction of enzyme and side chain of substrate.
authors.affiliation: Department of Biofunctional Chemistry, Graduate School of Natural Science and Technology, Okayama University, Japan.
authors.forename: Tomohiro
authors.initials: T
authors.lastname: Utsumi
authors.forename: Jiro
authors.initials: J
authors.lastname: Arima
authors.forename: Chika
authors.initials: C
authors.lastname: Sakaguchi
authors.forename: Takashi
authors.initials: T
authors.lastname: Tamura
authors.forename: Chiduko
authors.initials: C
authors.lastname: Sasaki
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Kusakabe
authors.forename: Shigetoshi
authors.initials: S
authors.lastname: Sugio
authors.forename: Kenji
authors.initials: K
authors.lastname: Inagaki
mesh_terms: D000594:Amino Acid Oxidoreductases / Q000737:chemistry* / Q000235:genetics; D001120:Arginine / Q000737:chemistry* / Q000235:genetics; D002384:Catalysis; D020134:Catalytic Domain; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D009154:Mutation; D011487:Protein Conformation; D013302:Streptomyces / Q000201:enzymology*; D013379:Substrate Specificity / Q000235:genetics
chemical_list: D001120:Arginine; D000594:Amino Acid Oxidoreductases; C023061:L-glutamate oxidase