title: Significant enhancement in the binding of p-nitrophenyl-beta-D-xylobioside by the E128H mutant F/10 xylanase from Streptomyces olivaceoviridis E-86.
abstract: Mutagenesis studies were carried out to examine the effects of replacement of either the nucleophile Glu-236 or the acid/base Glu-128 residue of the F/10 xylanase by a His residue. To our surprise, the affinity for the p-nitrophenyl-beta-D-xylobioside substrate was increased by 10(3)-fold in the case of the mutant E128H enzyme compared with that of the wild-type F/10 xylanase. The catalytic activity of the mutant enzymes was low, despite the fact that the distance between the nucleophilic atom (an oxygen in the native xylanase and a nitrogen in the mutant) and the alpha-carbon was barely changed. Thus, the alteration of the acid/base functionality (Glu-128 to His mutation) provided a significantly favorable interaction within the E128H enzyme/substrate complex in the ground state, accompanying a reduction in the stabilization effect in the transition state.
authors.affiliation: Institute of Applied Biochemistry, University of Tsukuba, Japan.
authors.forename: A
authors.initials: A
authors.lastname: Kuno
authors.forename: D
authors.initials: D
authors.lastname: Shimizu
authors.forename: S
authors.initials: S
authors.lastname: Kaneko
authors.forename: T
authors.initials: T
authors.lastname: Hasegawa
authors.forename: Y
authors.initials: Y
authors.lastname: Gama
authors.forename: K
authors.initials: K
authors.lastname: Hayashi
authors.forename: I
authors.initials: I
authors.lastname: Kusakabe
authors.forename: K
authors.initials: K
authors.lastname: Taira
mesh_terms: D004187:Disaccharides / Q000378:metabolism*; D043364:Endo-1,4-beta Xylanases; D018698:Glutamic Acid / Q000235:genetics / Q000378:metabolism; D006639:Histidine / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D013302:Streptomyces / Q000201:enzymology* / Q000235:genetics; D014995:Xylosidases / Q000235:genetics / Q000378:metabolism*
chemical_list: C100564:2-nitrophenylxylobioside; D004187:Disaccharides; D018698:Glutamic Acid; D006639:Histidine; D014995:Xylosidases; D043364:Endo-1,4-beta Xylanases