Transhalogenation Catalysed by Haloalkane Dehalogenases Engineered to Stop Natural Pathway at Intermediate

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F00159816%3A_____%2F19%3A00071029" target="_blank" >RIV/00159816:_____/19:00071029 - isvavai.cz</a>

Nalezeny alternativní kódy

RIV/00216224:14310/19:00113347

Výsledek na webu

<a href="https://onlinelibrary.wiley.com/doi/pdf/10.1002/adsc.201900132" target="_blank" >https://onlinelibrary.wiley.com/doi/pdf/10.1002/adsc.201900132</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1002/adsc.201900132" target="_blank" >10.1002/adsc.201900132</a>

Jazyk výsledku

angličtina

Název v původním jazyce

Transhalogenation Catalysed by Haloalkane Dehalogenases Engineered to Stop Natural Pathway at Intermediate

Popis výsledku v původním jazyce

Haloalkane dehalogenases (HLDs) are alpha/beta-hydrolases that convert halogenated compounds to their corresponding alcohols. The overall kinetic mechanism proceeds via four steps: (i) binding of halogenated substrate, (ii) bimolecular nucleophilic substitution (S(N)2) leading to the cleavage of a carbon-halogen bond and the formation of an alkyl-enzyme intermediate, (iii) nucleophilic addition of a water molecule resulting in the hydrolysis of the intermediate to the corresponding alcohol and (iv) release of the reaction products - an alcohol, a halide ion and a proton. Although, the overall reaction has been reported as irreversible, several kinetic evidences from previous studies suggest the reversibility of the first S(N)2 chemical step. To study this phenomenon, we have engineered HLDs to stop the catalytic cycle at the stage of the alkyl-enzyme intermediate. The ability of the intermediate to exchange halides was confirmed by a stopped-flow fluorescence binding analysis. Finally, the transhalogenation reaction was confirmed with several HLDs and 2,3-dichloropropene in the presence of a high concentration of iodide. The formation of the transhalogenation product 3-iodo-2-chloropropene catalysed by five mutant HLDs was identified by gas chromatography coupled with mass spectrometry. Hereby we demonstrated the reversibility of the cleavage of the carbon-halogen bond by HLDs resulting in a transhalogenation. After optimization, the transhalogenation reaction can possibly find its use in biocatalytic applications. Enabling this reaction by strategically engineering the enzyme to stop at an intermediate in the catalytic cycle that is synthetically more useful than the product of the natural pathway is a novel concept.

Název v anglickém jazyce

Transhalogenation Catalysed by Haloalkane Dehalogenases Engineered to Stop Natural Pathway at Intermediate

Popis výsledku anglicky

Haloalkane dehalogenases (HLDs) are alpha/beta-hydrolases that convert halogenated compounds to their corresponding alcohols. The overall kinetic mechanism proceeds via four steps: (i) binding of halogenated substrate, (ii) bimolecular nucleophilic substitution (S(N)2) leading to the cleavage of a carbon-halogen bond and the formation of an alkyl-enzyme intermediate, (iii) nucleophilic addition of a water molecule resulting in the hydrolysis of the intermediate to the corresponding alcohol and (iv) release of the reaction products - an alcohol, a halide ion and a proton. Although, the overall reaction has been reported as irreversible, several kinetic evidences from previous studies suggest the reversibility of the first S(N)2 chemical step. To study this phenomenon, we have engineered HLDs to stop the catalytic cycle at the stage of the alkyl-enzyme intermediate. The ability of the intermediate to exchange halides was confirmed by a stopped-flow fluorescence binding analysis. Finally, the transhalogenation reaction was confirmed with several HLDs and 2,3-dichloropropene in the presence of a high concentration of iodide. The formation of the transhalogenation product 3-iodo-2-chloropropene catalysed by five mutant HLDs was identified by gas chromatography coupled with mass spectrometry. Hereby we demonstrated the reversibility of the cleavage of the carbon-halogen bond by HLDs resulting in a transhalogenation. After optimization, the transhalogenation reaction can possibly find its use in biocatalytic applications. Enabling this reaction by strategically engineering the enzyme to stop at an intermediate in the catalytic cycle that is synthetically more useful than the product of the natural pathway is a novel concept.

Druh

J_imp - Článek v periodiku v databázi Web of Science

CEP obor

—

OECD FORD obor

10406 - Analytical chemistry

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)

Rok uplatnění

2019

Kód důvěrnosti údajů

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

Název periodika

ADVANCED SYNTHESIS &amp; CATALYSIS

ISSN

1615-4150

e-ISSN

—

Svazek periodika

361

Číslo periodika v rámci svazku

11

Stát vydavatele periodika

US - Spojené státy americké

Počet stran výsledku

5

Strana od-do

2438-2442

Kód UT WoS článku

000471070400007

EID výsledku v databázi Scopus

—