Ancestral Haloalkane Dehalogenases Show Robustness and Unique Substrate Specificity
Identifikátory výsledku
Kód výsledku v IS VaVaI
<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F00159816%3A_____%2F17%3A00067125" target="_blank" >RIV/00159816:_____/17:00067125 - isvavai.cz</a>
Nalezeny alternativní kódy
RIV/00216224:14310/17:00095408
Výsledek na webu
<a href="http://dx.doi.org/10.1002/cbic.201700197" target="_blank" >http://dx.doi.org/10.1002/cbic.201700197</a>
DOI - Digital Object Identifier
<a href="http://dx.doi.org/10.1002/cbic.201700197" target="_blank" >10.1002/cbic.201700197</a>
Alternativní jazyky
Jazyk výsledku
angličtina
Název v původním jazyce
Ancestral Haloalkane Dehalogenases Show Robustness and Unique Substrate Specificity
Popis výsledku v původním jazyce
Ancestral sequence reconstruction (ASR) represents a powerful approach for empirical testing structure-function relationships of diverse proteins. We employed ASR to predict sequences of five ancestral haloalkane dehalogenases (HLDs) from the HLD-II subfamily. Genes encoding the inferred ancestral sequences were synthesized and expressed in Escherichia coli, and the resurrected ancestral enzymes (AncHLD1-5) were experimentally characterized. Strikingly, the ancestral HLDs exhibited significantly enhanced thermodynamic stability compared to extant enzymes (Delta T-m up to 24 degrees C), as well as higher specific activities with preference for short multi-substituted halogenated substrates. Moreover, multivariate statistical analysis revealed a shift in the substrate specificity profiles of AncHLD1 and AncHLD2. This is extremely difficult to achieve by rational protein engineering. The study highlights that ASR is an efficient approach for the development of novel biocatalysts and robust templates for directed evolution.
Název v anglickém jazyce
Ancestral Haloalkane Dehalogenases Show Robustness and Unique Substrate Specificity
Popis výsledku anglicky
Ancestral sequence reconstruction (ASR) represents a powerful approach for empirical testing structure-function relationships of diverse proteins. We employed ASR to predict sequences of five ancestral haloalkane dehalogenases (HLDs) from the HLD-II subfamily. Genes encoding the inferred ancestral sequences were synthesized and expressed in Escherichia coli, and the resurrected ancestral enzymes (AncHLD1-5) were experimentally characterized. Strikingly, the ancestral HLDs exhibited significantly enhanced thermodynamic stability compared to extant enzymes (Delta T-m up to 24 degrees C), as well as higher specific activities with preference for short multi-substituted halogenated substrates. Moreover, multivariate statistical analysis revealed a shift in the substrate specificity profiles of AncHLD1 and AncHLD2. This is extremely difficult to achieve by rational protein engineering. The study highlights that ASR is an efficient approach for the development of novel biocatalysts and robust templates for directed evolution.
Klasifikace
Druh
J<sub>imp</sub> - Článek v periodiku v databázi Web of Science
CEP obor
—
OECD FORD obor
10608 - Biochemistry and molecular biology
Návaznosti výsledku
Projekt
Výsledek vznikl pri realizaci vícero projektů. Více informací v záložce Projekty.
Návaznosti
P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)
Ostatní
Rok uplatnění
2017
Kód důvěrnosti údajů
S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů
Údaje specifické pro druh výsledku
Název periodika
ChemBioChem
ISSN
1439-4227
e-ISSN
—
Svazek periodika
18
Číslo periodika v rámci svazku
14
Stát vydavatele periodika
DE - Spolková republika Německo
Počet stran výsledku
9
Strana od-do
1448-1456
Kód UT WoS článku
000405726100015
EID výsledku v databázi Scopus
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