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Significant enhancement in the binding of p ‐nitrophenyl‐β‐ d ‐xylobioside by the E128H mutant F/10 xylanase from Streptomyces olivaceoviridis E‐86
Author(s) -
Kuno Atsushi,
Shimizu Daisuke,
Kaneko Satoshi,
Hasegawa Tsunemi,
Gama Yasuo,
Hayashi Kiyoshi,
Kusakabe Isao,
Taira Kazunari
Publication year - 1999
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/s0014-5793(99)00498-6
Subject(s) - xylanase , mutant , chemistry , stereochemistry , nucleophile , enzyme , substrate (aquarium) , residue (chemistry) , site directed mutagenesis , mutagenesis , biochemistry , wild type , catalysis , biology , gene , ecology
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‐β‐ 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 α‐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.