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Engineering endoglucanase II from Trichoderma reesei to improve the catalytic efficiency at a higher pH optimum

Paper ID Volume ID Publish Year Pages File Format Full-Text
24827 43539 2008 6 PDF Available
Title
Engineering endoglucanase II from Trichoderma reesei to improve the catalytic efficiency at a higher pH optimum
Abstract

The catalytic efficiency and pH optimum of Trichoderma reesei endo-β-1,4-glucanase II were improved by protein engineering. We subjected residue 342 to saturation mutagenesis, and further changed the enzyme by random mutagenesis and two rounds of DNA shuffling. Enzyme variants were purified and characterized. Variant N342V exhibited an optimal activity at pH 5.8, corresponding to a basic shift of 1 pH unit compared with the wild-type enzyme, and had improved catalytic efficiency (1.5-fold of kcat/Km) for the main substrates at pH 6.2. Variants N342R and N39R/L218H/W276R/N342T both had a pH optimum of 6.2 and the latter had improved catalytic efficiency (1.4-fold of kcat/Km) at pH 6.2. Variants L218H, Q139R/N342T and Q139R/L218H/W276R/N342T all had more than 4.5-fold higher activity in reactions compared with the wild-type at pH 7.0. The relationship between the structures and the activities of the variants were analyzed by modeling the structures of the endoglucanase II variants. More stable helixes and changed electrostatic interactions between the catalytic residues and substrates may explain the higher activities and higher pH optima of the variants.

Keywords
Endoglucanase II; Trichoderma reesei; Saturation mutation; DNA shuffling; pH optimum; Catalytic efficiency
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Engineering endoglucanase II from Trichoderma reesei to improve the catalytic efficiency at a higher pH optimum
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 135, Issue 2, 1 June 2008, Pages 190–195
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us