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Amino acid residues adjacent to the catalytic cavity of tetramer l-asparaginase II contribute significantly to its catalytic efficiency and thermostability

Paper ID Volume ID Publish Year Pages File Format Full-Text
16891 42621 2016 8 PDF Available
Title
Amino acid residues adjacent to the catalytic cavity of tetramer l-asparaginase II contribute significantly to its catalytic efficiency and thermostability
Abstract

•Site directed mutagenesis was applied to increase thermostability and enzyme activity of BsAII.•One mutant showed 83.10% and 17.5 °C increase in specific activity and half-inactivation temperature, respectively.•The structure changes might contribute to the improved activity.•The increased hydrophilicity and surface charge redistribution helped stabilize the conformation of the enzyme.

l-Asparaginase (l-asparagine amidohydrolase, EC 3.5.1.1) catalyzes the hydrolysis of l-asparagine to l-aspartic acid and ammonia. It can be used to reduce the formation of acrylamide, which is carcinogenic to humans in foods, via removal of the precursor, asparagine, from the primary ingredients. However, low activity and poor thermostability of l-asparaginase restrict its application in food industry. In this study, we successfully improved thermostability and catalytic efficiency of l-asparaginase II (BsAII) from Bacillus subtilis B11-06 by site-directed mutagenesis. According to sequences alignment and homologous modeling, residues G107, T109 and S166 which were adjacent to the catalytic cavity were selected and substituted by Asp, Gln/Ser and Ala, respectively, to construct mutants G107D, T109Q, T109S and S166A. The BsAII mutant of G107D (G107Dansz) displayed superior performance in thermal tolerance and higher activity than the wild-type enzyme (towards l-asparagine). Comparative analysis of hydrogen bond interactions, surface electrostatic potential and structure of substrate binding pocket between G107Danszand BsAII indicated that the substitution of G107, which was adjacent to catalytic cavity with Asp, resulted in small conformational changes and surface electrostatic potential redistribution and contributed to the improved protein stability and catalytic efficiency.

Keywords
l-Asparaginase II; Homologous modeling; Site-directed mutagenesis; Thermostability; Enzyme activity
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Amino acid residues adjacent to the catalytic cavity of tetramer l-asparaginase II contribute significantly to its catalytic efficiency and thermostability
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Publisher
Database: Elsevier - ScienceDirect
Journal: Enzyme and Microbial Technology - Volume 82, January 2016, Pages 15–22
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