Affiliation:
1. Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University, Kyoto Japan
2. Department of Life Science and Technology, Tokyo Institute of Technology, Midori-ku, Japan
Abstract
ABSTRACT
Site saturation mutagenesis library is a recently developed technique, in which any one out of all amino acid residues in a target region is substituted into other 19 amino acid residues. In this study, we used this technique to increase the thermostability of a GH10 xylanase, XynR, from Bacillus sp. strain TAR-1. We hypothesized that the substrate binding region of XynR is flexible, and that the thermostability of XynR will increase if the flexibility of the substrate binding region is decreased without impairing the substrate binding ability. Site saturation mutagenesis libraries of amino acid residues Tyr43–Lys115 and Ala300–Asn325 of XynR were constructed. By screening 480 clones, S92E was selected as the most thermostable one, exhibiting the residual activity of 80% after heat treatment at 80°C for 15 min in the hydrolysis of Remazol Brilliant Blue-xylan. Our results suggest that this strategy is effective for stabilization of GH10 xylanase.
Abbreviations: DNS: 3,5-dinitrosalicylic acid; RBB-xylan: Remazol Brilliant Blue-xylan
Funder
Japan Science and Technology Agency
Japan Society for the Promotion of Science
Publisher
Oxford University Press (OUP)
Subject
Organic Chemistry,Molecular Biology,Applied Microbiology and Biotechnology,General Medicine,Biochemistry,Analytical Chemistry,Biotechnology
Cited by
10 articles.
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