Deciphering Enzyme Mechanisms with Engineered Ancestors and Substrate Analogues

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F00159816%3A_____%2F23%3A00079702" target="_blank" >RIV/00159816:_____/23:00079702 - isvavai.cz</a>

Nalezeny alternativní kódy

RIV/00216224:14310/23:00132027

Výsledek na webu

<a href="https://chemistry-europe.onlinelibrary.wiley.com/doi/10.1002/cctc.202300745" target="_blank" >https://chemistry-europe.onlinelibrary.wiley.com/doi/10.1002/cctc.202300745</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1002/cctc.202300745" target="_blank" >10.1002/cctc.202300745</a>

Jazyk výsledku

angličtina

Název v původním jazyce

Deciphering Enzyme Mechanisms with Engineered Ancestors and Substrate Analogues

Popis výsledku v původním jazyce

Environmentally friendly industrial and biotech processes greatly benefit from enzyme-based technologies. Their use is often possible only when the enzyme-catalytic mechanism is thoroughly known. Thus, atomic-level knowledge of a Michaelis enzyme-substrate complex, revealing molecular details of substrate recognition and catalytic chemistry, is crucial for understanding and then rationally extending or improving enzyme-catalyzed reactions. However, many known enzymes sample huge protein conformational space, often preventing complete structural characterization by X-ray crystallography. Moreover, using a cognate substrate is problematic since its conversion into a reaction product in the presence of the enzyme will prevent the capture of the enzyme-substrate conformation in an activated state. Here, we outlined how to deal with such obstacles, focusing on the recent discovery of a Renilla-type bioluminescence reaction mechanism facilitated by a combination of engineered ancestral enzyme and the availability of a non-oxidizable luciferin analogue. The automated ancestral sequence reconstructions using FireProtASR provided a thermostable enzyme suited for structural studies, and a stable luciferin analogue azacoelenterazine provided a structurally cognate chemical incapable of catalyzed oxidation. We suggest that an analogous strategy can be applied to various enzymes with unknown catalytic mechanisms and poor crystallizability. Many known enzymes sample huge protein conformational space, hampering structural characterization by X-ray crystallography, and preventing thus the understanding of their catalytic mechanisms. In this review, we outline that the combination of reconstructed ancestral enzymes with unconvertible substrate analogues is becoming a powerful strategy to decipher the challenging mechanisms of enzyme catalysis.image

Název v anglickém jazyce

Deciphering Enzyme Mechanisms with Engineered Ancestors and Substrate Analogues

Popis výsledku anglicky

Environmentally friendly industrial and biotech processes greatly benefit from enzyme-based technologies. Their use is often possible only when the enzyme-catalytic mechanism is thoroughly known. Thus, atomic-level knowledge of a Michaelis enzyme-substrate complex, revealing molecular details of substrate recognition and catalytic chemistry, is crucial for understanding and then rationally extending or improving enzyme-catalyzed reactions. However, many known enzymes sample huge protein conformational space, often preventing complete structural characterization by X-ray crystallography. Moreover, using a cognate substrate is problematic since its conversion into a reaction product in the presence of the enzyme will prevent the capture of the enzyme-substrate conformation in an activated state. Here, we outlined how to deal with such obstacles, focusing on the recent discovery of a Renilla-type bioluminescence reaction mechanism facilitated by a combination of engineered ancestral enzyme and the availability of a non-oxidizable luciferin analogue. The automated ancestral sequence reconstructions using FireProtASR provided a thermostable enzyme suited for structural studies, and a stable luciferin analogue azacoelenterazine provided a structurally cognate chemical incapable of catalyzed oxidation. We suggest that an analogous strategy can be applied to various enzymes with unknown catalytic mechanisms and poor crystallizability. Many known enzymes sample huge protein conformational space, hampering structural characterization by X-ray crystallography, and preventing thus the understanding of their catalytic mechanisms. In this review, we outline that the combination of reconstructed ancestral enzymes with unconvertible substrate analogues is becoming a powerful strategy to decipher the challenging mechanisms of enzyme catalysis.image

Druh

J_imp - Článek v periodiku v databázi Web of Science

CEP obor

—

OECD FORD obor

10403 - Physical chemistry

Projekt

Výsledek vznikl pri realizaci vícero projektů. Více informací v záložce Projekty.

Návaznosti

—

Rok uplatnění

2023

Kód důvěrnosti údajů

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

Název periodika

ChemCatChem

ISSN

1867-3880

e-ISSN

1867-3899

Svazek periodika

15

Číslo periodika v rámci svazku

19

Stát vydavatele periodika

DE - Spolková republika Německo

Počet stran výsledku

15

Strana od-do

—

Kód UT WoS článku

001060401600001

EID výsledku v databázi Scopus

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