Comparative analysis of point mutations on protein COOH terminal near surface and its hydrophobic core provide insights on thermostability of Bacillus Lipase LipJ
{"title":"Comparative analysis of point mutations on protein COOH terminal near surface and its hydrophobic core provide insights on thermostability of Bacillus Lipase LipJ","authors":"Shelly Goomber, Nisha Chopra, Gursimran Kaur Bedi, Jagdeep Kaur","doi":"10.1016/j.molcatb.2017.04.001","DOIUrl":null,"url":null,"abstract":"<div><p>Amino acid composition, protein conformation, weak interactions are some of the factors determining protein stability. Two point mutants of <em>Bacillus</em> LipJ (Ala81Tyr and Asn166Tyr) were designed and generated for comparison. Residue Ala81 targeted for substitution was buried in protein core with 0% accessibility. On other hand, residue Asn166 at protein <img>COOH terminal was predicted to be located within αF helix near surface. Point variants Ala81Tyr and Asn166Tyr were generated by site directed mutagenesis. In comparison to native protein LipJ, point variant Ala81Tyr was observed to be more thermostable and demonstrated 1.6 times relative activity after 1<!--> <!-->h exposure at 50<!--> <!-->°C. However, point variant Asn166Tyr showed dramatic shift in its thermostability, specific activity and substrate specificity pattern compared to wild type LipJ. Asn166Tyr variant was found to remain stable for hours at 50<!--> <!-->°C. Specific activity of Asn166Tyr was 20% of parent enzyme. Substrate specificity of Asn166Tyr was significantly shifted to fatty acyl chains of shorter lengths. Homolog modelling predicted global vs local structure change to explain molecular basis of differential behavior of point variants Asn166Tyr and Ala81Tyr respectively.</p></div>","PeriodicalId":16416,"journal":{"name":"Journal of Molecular Catalysis B-enzymatic","volume":"133 ","pages":"Pages S482-S490"},"PeriodicalIF":0.0000,"publicationDate":"2016-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/j.molcatb.2017.04.001","citationCount":"1","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Journal of Molecular Catalysis B-enzymatic","FirstCategoryId":"1085","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S1381117717300450","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"Chemical Engineering","Score":null,"Total":0}
引用次数: 1
Abstract
Amino acid composition, protein conformation, weak interactions are some of the factors determining protein stability. Two point mutants of Bacillus LipJ (Ala81Tyr and Asn166Tyr) were designed and generated for comparison. Residue Ala81 targeted for substitution was buried in protein core with 0% accessibility. On other hand, residue Asn166 at protein COOH terminal was predicted to be located within αF helix near surface. Point variants Ala81Tyr and Asn166Tyr were generated by site directed mutagenesis. In comparison to native protein LipJ, point variant Ala81Tyr was observed to be more thermostable and demonstrated 1.6 times relative activity after 1 h exposure at 50 °C. However, point variant Asn166Tyr showed dramatic shift in its thermostability, specific activity and substrate specificity pattern compared to wild type LipJ. Asn166Tyr variant was found to remain stable for hours at 50 °C. Specific activity of Asn166Tyr was 20% of parent enzyme. Substrate specificity of Asn166Tyr was significantly shifted to fatty acyl chains of shorter lengths. Homolog modelling predicted global vs local structure change to explain molecular basis of differential behavior of point variants Asn166Tyr and Ala81Tyr respectively.
期刊介绍：
Journal of Molecular Catalysis B: Enzymatic is an international forum for researchers and product developers in the applications of whole-cell and cell-free enzymes as catalysts in organic synthesis. Emphasis is on mechanistic and synthetic aspects of the biocatalytic transformation.
Papers should report novel and significant advances in one or more of the following topics;
Applied and fundamental studies of enzymes used for biocatalysis;
Industrial applications of enzymatic processes, e.g. in fine chemical synthesis;
Chemo-, regio- and enantioselective transformations;
Screening for biocatalysts;
Integration of biocatalytic and chemical steps in organic syntheses;
Novel biocatalysts, e.g. enzymes from extremophiles and catalytic antibodies;
Enzyme immobilization and stabilization, particularly in non-conventional media;
Bioprocess engineering aspects, e.g. membrane bioreactors;
Improvement of catalytic performance of enzymes, e.g. by protein engineering or chemical modification;
Structural studies, including computer simulation, relating to substrate specificity and reaction selectivity;
Biomimetic studies related to enzymatic transformations.
求助内容：
应助结果提醒方式：
京公网安备 11010802042870号
