Academic Journal
Structural Characterization and Directed Evolution of a Novel Acetyl Xylan Esterase Reveals Thermostability Determinants of the Carbohydrate Esterase 7 Family
| Title: | Structural Characterization and Directed Evolution of a Novel Acetyl Xylan Esterase Reveals Thermostability Determinants of the Carbohydrate Esterase 7 Family |
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| Authors: | Adesioye, Fiyinfoluwa A., Makhalanyane, Thulani P., Vikram, Surendra, Sewell, Bryan T., Schubert, Wolf-Dieter, Cowan, Don A. |
| Publisher Information: | American Society for Microbiology |
| Publication Year: | 2018 |
| Collection: | PubMed Central (PMC) |
| Subject Terms: | Enzymology and Protein Engineering |
| Description: | A hot desert hypolith metagenomic DNA sequence data set was screened in silico for genes annotated as acetyl xylan esterases (AcXEs). One of the genes identified encoded an ∼36-kDa protein (Axe1NaM1). The synthesized gene was cloned and expressed, and the resulting protein was purified. NaM1 was optimally active at pH 8.5 and 30°C and functionally stable at salt concentrations of up to 5 M. The specific activity and catalytic efficiency were 488.9 U mg−1 and 3.26 × 106 M−1 s−1, respectively. The crystal structure of wild-type NaM1 was solved at a resolution of 2.03 Å, and a comparison with the structures and models of more thermostable carbohydrate esterase 7 (CE7) family enzymes and variants of NaM1 from a directed evolution experiment suggests that reduced side-chain volume of protein core residues is relevant to the thermal stability of NaM1. Surprisingly, a single point mutation (N96S) not only resulted in a simultaneous improvement in thermal stability and catalytic efficiency but also increased the acyl moiety substrate range of NaM1. |
| Document Type: | text |
| Language: | English |
| Relation: | http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5881061/; http://www.ncbi.nlm.nih.gov/pubmed/29453256; http://dx.doi.org/10.1128/AEM.02695-17 |
| DOI: | 10.1128/AEM.02695-17 |
| Availability: | https://doi.org/10.1128/AEM.02695-17 http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5881061/ http://www.ncbi.nlm.nih.gov/pubmed/29453256 |
| Rights: | Copyright © 2018 American Society for Microbiology. ; https://doi.org/10.1128/ASMCopyrightv2 ; All Rights Reserved (https://doi.org/10.1128/ASMCopyrightv2) . |
| Accession Number: | edsbas.70F5F80C |
| Database: | BASE |
| Description not available. |