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Virtual screening of mandelate racemase mutants with enhanced activity based on binding energy in the transition state

Paper ID Volume ID Publish Year Pages File Format Full-Text
17239 42653 2014 7 PDF Available
Title
Virtual screening of mandelate racemase mutants with enhanced activity based on binding energy in the transition state
Abstract

•A virtual screening method based on the binding energy was developed.•The method could efficiently predict the mutational effect on catalysis.•Mutants exhibiting enhanced activities to non-natural substrates were obtained.•Enhanced interaction between the active site and substrate led to higher activity.

Mandelate racemase (MR) is a promising candidate for the dynamic kinetic resolution of racemates. However, the poor activity of MR towards most of its non-natural substrates limits its widespread application. In this work, a virtual screening method based on the binding energy in the transition state was established to assist in the screening of MR mutants with enhanced catalytic efficiency. Using R-3-chloromandelic acid as a model substrate, a total of 53 mutants were constructed based on rational design in the two rounds of screening. The number of mutants for experimental validation was brought down to 17 by the virtual screening method, among which 14 variants turned out to possess improved catalytic efficiency. The variant V26I/Y54V showed 5.2-fold higher catalytic efficiency (kcat/Km) towards R-3-chloromandelic acid than that observed for the wild-type enzyme. Using this strategy, mutants were successfully obtained for two other substrates, R-mandelamide and R-2-naphthylglycolate (V26I and V29L, respectively), both with a 2-fold improvement in catalytic efficiency. These results demonstrated that this method could effectively predict the trend of mutational effects on catalysis. Analysis from the energetic and structural assays indicated that the enhanced interactions between the active sites and the substrate in the transition state led to improved catalytic efficiency. It was concluded that this virtual screening method based on the binding energy in the transition state was beneficial in enzyme rational redesign and helped to better understand the catalytic properties of the enzyme.

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Keywords
Virtual screening; Mandelate racemase; Binding energy; Transition state; Molecular dynamics simulation
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Virtual screening of mandelate racemase mutants with enhanced activity based on binding energy in the transition state
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Publisher
Database: Elsevier - ScienceDirect
Journal: Enzyme and Microbial Technology - Volume 55, 5 February 2014, Pages 121–127
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