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Engineering a thermostable β-1,3-1,4-glucanase from Paecilomyces thermophila to improve catalytic efficiency at acidic pH

Paper ID Volume ID Publish Year Pages File Format Full-Text
23691 43463 2012 6 PDF Available
Title
Engineering a thermostable β-1,3-1,4-glucanase from Paecilomyces thermophila to improve catalytic efficiency at acidic pH
Abstract

To fulfill the need for acid-tolerant and thermostable β-1,3-1,4-glucanases, an error-prone PCR and DNA-shuffling approach was employed to enhance the activity of thermostable β-1,3-1,4-glucanases from Paecilomyces thermophila (PtLic16A) at acidic pH. Mutant PtLic16AM2 was selected and characterized, and showed optimal activity at pH 5.0, corresponding to an acidic shift of 2.0 pH units relative to the wild-type enzyme. Other properties of PtLic16A such as temperature optimum and substrate specificity that are beneficial for industrial applications did not change. Based on the substituted residues of PtLic16AM2, three site-directed mutations, D56G, D221G and C263S, were designed to study these residues’ roles. The amino acid residues at positions 56 and 263 were found to be important in determining optimal pH activity. Activity of the D221G variant showed no significant difference from the wild-type. Thus, it appears that the change in optimal pH for PtLic16AM2 was mainly caused by the combination of substitutions D56G and C263S. This study provides a β-1,3-1,4-glucanase (PtLic16AM2) with high potential for industrial applications.

► The activity of thermostable β-1,3-1,4-glucanases was enhanced at acidic pH. ► The error-prone PCR and DNA shuffling approach was employed for directed evolution. ► The optimal pH of mutant PtLic16AM2 was shifted from 7.0 to 5.0. ► The change of optimal pH of PtLic16AM2 was mainly caused by D56G and C263S together.

Keywords
β-1,3-1,4-Glucanase; Directed evolution; Optimal pH; Site-directed mutagenesis; Paecilomyces thermophila
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Engineering a thermostable β-1,3-1,4-glucanase from Paecilomyces thermophila to improve catalytic efficiency at acidic pH
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 159, Issues 1–2, 31 May 2012, Pages 50–55
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