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Tailor-made β-glucosidase with increased activity at lower temperature without loss of stability and glucose tolerance

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Abstract

The hydrolysis of cellobiose by β-glucosidases represents the crucial step in the degradation of lignocellulosic biomass and is therefore a major target for improvement by enzyme engineering. BglA from Caldicellulosiruptor saccharolyticus (CsBglA) is among the β-glucosidases with highest known activity and thermostability, however with a temperature optimum of 70–80 °C. To improve the enzyme's activity at moderate temperatures, a semi-rational approach based on homology modeling, conservation scoring and subsequent site-saturation mutagenesis was combined with random mutagenesis, resulting in the variant CsBglA-LYTH with 150% improvement of kcat/KM at 55 °C, a specific activity of 504 U mg−1 (CsBglA: 359 U mg−1) and a KM-value of 37.1 mM (CsBglA: 61.2 mM). The temperature optimum of the enzyme was altered from 80 °C to 65 °C while retaining stability at the temperature of 55 °C as applied for cellulose hydrolysis. To our knowledge, this makes CsBglA-LYTH the bacterial β-glucosidase with highest activity and stability.

Graphical abstract: Tailor-made β-glucosidase with increased activity at lower temperature without loss of stability and glucose tolerance

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Supplementary files

Article information


Submitted
05 Dec 2019
Accepted
11 Mar 2020
First published
11 Mar 2020

Green Chem., 2020, Advance Article
Article type
Paper

Tailor-made β-glucosidase with increased activity at lower temperature without loss of stability and glucose tolerance

F. Lenz, P. Zurek, M. Umlauf, I. E. P. Tozakidis and J. Jose, Green Chem., 2020, Advance Article , DOI: 10.1039/C9GC04166D

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