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Title: Engineering glycoside hydrolase stability by the introduction of zinc binding

The development of robust enzymes, in particular cellulases, is a key step in the success of biological routes to `second-generation' biofuels. The typical sources of the enzymes used to degrade biomass include mesophilic and thermophilic organisms. The endoglucanase J30 from glycoside hydrolase family 9 was originally identified through metagenomic analyses of compost-derived bacterial consortia. These studies, which were tailored to favor growth on targeted feedstocks, have already been shown to identify cellulases with considerable thermal tolerance. The amino-acid sequence of J30 shows comparably low identity to those of previously analyzed enzymes. As an enzyme that combines a well measurable activity with a relatively low optimal temperature (50°C) and a modest thermal tolerance, it offers the potential for structural optimization aimed at increased stability. Here, the crystal structure of wild-type J30 is presented along with that of a designed triple-mutant variant with improved characteristics for industrial applications. Through the introduction of a structural Zn 2+ site, the thermal tolerance was increased by more than 10°C and was paralleled by an increase in the catalytic optimum temperature by more than 5°C.
Authors:
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Publication Date:
Grant/Contract Number:
AC02-76SF00515
Type:
Published Article
Journal Name:
Acta Crystallographica Section D Structural Biology
Additional Journal Information:
Journal Volume: 74; Journal Issue: 7; Related Information: CHORUS Timestamp: 2018-06-29 11:40:12; Journal ID: ISSN 2059-7983
Publisher:
International Union of Crystallography (IUCr)
Sponsoring Org:
USDOE
Country of Publication:
United Kingdom
Language:
English
OSTI Identifier:
1457487