Engineering glycoside hydrolase stability by the introduction of zinc binding

Ellinghaus, Thomas L.; Pereira, Jose H.; McAndrew, Ryan P.; Welner, Ditte H.; DeGiovanni, Andy M.; Guenther, Joel M.; Tran, Huu M.; Feldman, Taya; Simmons, Blake A.; Sale, Kenneth L.; Adams, Paul D.

doi:10.1107/S2059798318006678

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

Journal Article · 27 June 2018 · Acta Crystallographica. Section D. Structural Biology

DOI: https://doi.org/10.1107/S2059798318006678 · OSTI ID:1457487

Ellinghaus, Thomas L. ^[1]; Pereira, Jose H. ^[1]; McAndrew, Ryan P. ^[1];

^[1]; DeGiovanni, Andy M. ^[1]; Guenther, Joel M. ^[2]; Tran, Huu M. ^[2]; Feldman, Taya ^[2]; Simmons, Blake A. ^[3]; Sale, Kenneth L. ^[2]; Adams, Paul D. ^[4]

Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Molecular Biophysics and Integrated Bioimaging Division
Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Sandia National Lab. (SNL-CA), Livermore, CA (United States). Biological and Engineering Sciences Center
Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Biological Systems and Engineering Division
Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Molecular Biophysics and Integrated Bioimaging Division; Univ. of California, Berkeley, CA (United States). Dept. of Bioengineering

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 Zn²⁺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.

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Research Organization:: Lawrence Berkeley National Laboratory (LBNL), Berkeley, CA (United States); Sandia National Lab. (SNL-CA), Livermore, CA (United States)

Sponsoring Organization:: USDOE Office of Science (SC), Biological and Environmental Research (BER); National Institutes of Health (NIH); USDOE Office of Science (SC), Basic Energy Sciences (BES); USDOE

Grant/Contract Number:: AC02-05CH11231; AC02-76SF00515; AC04-94AL85000

OSTI ID:: 1457487

Alternate ID(s):: OSTI ID: 1476933; OSTI ID: 1506325

Report Number(s):: SAND-2018-9901J; ACSDAD; ark:/13030/qt2hd2g8vs

Journal Information:: Acta Crystallographica. Section D. Structural Biology, Vol. 74, Issue 7; ISSN 2059-7983

Publisher:: IUCrCopyright Statement

Country of Publication:: United States

Language:: English

Citation Metrics:

Cited by: 1 work

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