Exploration of Enzyme Diversity by Integrating Bioinformatics with Expression Analysis and Biochemical Characterization

Result code in IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F00216224%3A14310%2F18%3A00101756" target="_blank" >RIV/00216224:14310/18:00101756 - isvavai.cz</a>

Alternative codes found

RIV/00159816:_____/18:00068622

Result on the web

<a href="http://dx.doi.org/10.1021/acscatal.7b03523" target="_blank" >http://dx.doi.org/10.1021/acscatal.7b03523</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1021/acscatal.7b03523" target="_blank" >10.1021/acscatal.7b03523</a>

Result language

angličtina

Original language name

Exploration of Enzyme Diversity by Integrating Bioinformatics with Expression Analysis and Biochemical Characterization

Original language description

Millions of protein sequences are being discovered at an incredible pace, representing an inexhaustible source of biocatalysts. Here, we describe an integrated system for automated in silico screening and systematic characterization of diverse family members. The workflow consists of (i) identification and computational characterization of relevant genes by sequence/structural bioinformatics, (ii) expression analysis and activity screening of selected proteins, and (iii) complete biochemical/biophysical characterization and was validated against the haloalkane dehalogenase family. The sequence-based search identified 658 potential dehalogenases. The subsequent structural bioinformatics prioritized and selected 20 candidates for exploration of protein functional diversity. Out of these 20, the expression analysis and the robotic screening of enzymatic activity provided 8 soluble proteins with dehalogenase activity. The enzymes discovered originated from genetically unrelated Bacteria, Eukaryota, and also Archaea. Overall, the integrated system provided biocatalysts with broad catalytic diversity showing unique substrate specificity profiles, covering a wide range of optimal operational temperature from 20 to 70 degrees C and an unusually broad pH range from 5.7 to 10. We obtained the most catalytically proficient native haloalkane dehalogenase enzyme to date (k(cat)/K-0.5 = 96.8 mM(-1) s(-1) the most thermostable enzyme with melting temperature 71 degrees C, three different cold-adapted enzymes showing dehalogenase activity at near-to-zero temperatures, and a biocatalyst degrading the warfare chemical sulfur mustard. The established strategy can be adapted to other enzyme families for exploration of their biocatalytic diversity in a large sequence space continuously growing due to the use of next-generation sequencing technologies.

Czech name

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Czech description

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Type

J_imp - Article in a specialist periodical, which is included in the Web of Science database

CEP classification

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OECD FORD branch

10608 - Biochemistry and molecular biology

Project

Result was created during the realization of more than one project. More information in the Projects tab.

Continuities

P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)

Publication year

2018

Confidentiality

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Name of the periodical

ACS Catalysis

ISSN

2155-5435

e-ISSN

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Volume of the periodical

8

Issue of the periodical within the volume

3

Country of publishing house

US - UNITED STATES

Number of pages

21

Pages from-to

2402-2412

UT code for WoS article

000426804100087

EID of the result in the Scopus database

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