Exploring mechanism of enzyme catalysis by on-chip transient
kinetics coupled with global data analysis and molecular
modeling

J 2021

Exploring mechanism of enzyme catalysis by on-chip transient kinetics coupled with global data analysis and molecular modeling

HESS, David; Veronika DOČKALOVÁ; Piia Pauliina KOKKONEN; David BEDNÁŘ; Jiří DAMBORSKÝ et. al.

Základní údaje

Originální název

Exploring mechanism of enzyme catalysis by on-chip transient kinetics coupled with global data analysis and molecular modeling

Autoři

HESS, David; Veronika DOČKALOVÁ; Piia Pauliina KOKKONEN; David BEDNÁŘ; Jiří DAMBORSKÝ; Andrew DEMELLO; Zbyněk PROKOP a Stavros STAVRAKIS

Vydání

Chem, Cambridge, Cell Press, 2021, 2451-9294

Další údaje

Jazyk

angličtina

Typ výsledku

Článek v odborném periodiku

Obor

10400 1.4 Chemical sciences

Stát vydavatele

Spojené státy

Utajení

není předmětem státního či obchodního tajemství

Odkazy

URL

Impakt faktor

Impact factor: 25.832

Kód RIV

RIV/00216224:14310/21:00122285

Organizační jednotka

Přírodovědecká fakulta

DOI

https://doi.org/10.1016/j.chempr.2021.02.011

UT WoS

000652330200020

EID Scopus

2-s2.0-85103644244

Klíčová slova anglicky

HALOALKANE DEHALOGENASES; MICROFLUIDIC DEVICES; WATER; BIOCATALYSIS; DROPLETS; ENTROPY; BINDING

Štítky

14314010, 143180, H23, rivok

Příznaky

Mezinárodní význam, Recenzováno

Změněno: 15. 2. 2023 23:26, Mgr. Michaela Hylsová, Ph.D.

Anotace

V originále

The ability to engineer enzymes for industrial and biomedical applications is primarily limited by a paucity of mechanistic understanding. To gain insight into the mechanisms of enzyme catalysis, one must screen enormous numbers of discrete reaction conditions, which is a laborious task using conventional technologies. To address such limitations, we develop a droplet-based microfluidic platform for high-throughput acquisition of transient kinetic data over a range of substrate concentrations and temperatures. When compared with conventional methods, our platform reduces assay volumes by six orders of magnitude and increases throughput to 9,000 reactions/min. To demonstrate their utility, we measure the transient kinetics of three model enzymes, namely, beta-galactosidase, horseradish peroxidase, and microperoxidase. Additionally, we conduct a complex kinetic and thermodynamic study of engineered variants of haloalkane dehalogenases. Datasets are globally analyzed and complemented by molecular dynamics simulations, providing new insights into the molecular basis of substrate specificity and the role of hydration-related entropy.

Návaznosti

EF17_043/0009632, projekt VaV

Název: CETOCOEN Excellence

EF17_050/0008496, projekt VaV

Název: MSCAfellow@MUNI

LM2018121, projekt VaV

Název: Výzkumná infrastruktura RECETOX (Akronym: RECETOX RI)

Investor: Ministerstvo školství, mládeže a tělovýchovy ČR, RECETOX RI

857560, interní kód MU
(Kód CEP: EF17_043/0009632)

Název: CETOCOEN Excellence (Akronym: CETOCOEN Excellence)

Investor: Evropská unie, CETOCOEN Excellence, Spreading excellence and widening participation

Citovat

HESS, David; Veronika DOČKALOVÁ; Piia Pauliina KOKKONEN; David BEDNÁŘ; Jiří DAMBORSKÝ; Andrew DEMELLO; Zbyněk PROKOP a Stavros STAVRAKIS. Exploring mechanism of enzyme catalysis by on-chip transient kinetics coupled with global data analysis and molecular modeling. Chem. Cambridge: Cell Press, 2021, roč. 7, č. 4, s. 1066-1079. ISSN 2451-9294. Dostupné z: https://doi.org/10.1016/j.chempr.2021.02.011.

@article{1790417,
   author = {Hess, David and Dočkalová, Veronika and Kokkonen, Piia Pauliina and Bednář, David and Damborský, Jiří and DeMello, Andrew and Prokop, Zbyněk and Stavrakis, Stavros},
   article_location = {Cambridge},
   article_number = {4},
   doi = {https://doi.org/10.1016/j.chempr.2021.02.011},
   keywords = {HALOALKANE DEHALOGENASES; MICROFLUIDIC DEVICES; WATER; BIOCATALYSIS; DROPLETS; ENTROPY; BINDING},
   language = {eng},
   issn = {2451-9294},
   journal = {Chem},
   title = {Exploring mechanism of enzyme catalysis by on-chip transient kinetics coupled with global data analysis and molecular modeling},
   url = {https://www.sciencedirect.com/science/article/abs/pii/S2451929421000917?via%3Dihub},
   volume = {7},
   year = {2021}
}

TY  - JOUR
ID  - 1790417
AU  - Hess, David - Dočkalová, Veronika - Kokkonen, Piia Pauliina - Bednář, David - Damborský, Jiří - DeMello, Andrew - Prokop, Zbyněk - Stavrakis, Stavros
PY  - 2021
TI  - Exploring mechanism of enzyme catalysis by on-chip transient kinetics coupled with global data analysis and molecular modeling
JF  - Chem
VL  - 7
IS  - 4
SP  - 1066-1079
EP  - 1066-1079
PB  - Cell Press
SN  - 24519294
KW  - HALOALKANE DEHALOGENASES
KW  - MICROFLUIDIC DEVICES
KW  - WATER
KW  - BIOCATALYSIS
KW  - DROPLETS
KW  - ENTROPY
KW  - BINDING
UR  - https://www.sciencedirect.com/science/article/abs/pii/S2451929421000917?via%3Dihub
N2  - The ability to engineer enzymes for industrial and biomedical applications is primarily limited by a paucity of mechanistic understanding. To gain insight into the mechanisms of enzyme catalysis, one must screen enormous numbers of discrete reaction conditions, which is a laborious task using conventional technologies. To address such limitations, we develop a droplet-based microfluidic platform for high-throughput acquisition of transient kinetic data over a range of substrate concentrations and temperatures. When compared with conventional methods, our platform reduces assay volumes by six orders of magnitude and increases throughput to 9,000 reactions/min. To demonstrate their utility, we measure the transient kinetics of three model enzymes, namely, beta-galactosidase, horseradish peroxidase, and microperoxidase. Additionally, we conduct a complex kinetic and thermodynamic study of engineered variants of haloalkane dehalogenases. Datasets are globally analyzed and complemented by molecular dynamics simulations, providing new insights into the molecular basis of substrate specificity and the role of hydration-related entropy.
ER  -

HESS, David; Veronika DOČKALOVÁ; Piia Pauliina KOKKONEN; David BEDNÁŘ; Jiří DAMBORSKÝ; Andrew DEMELLO; Zbyněk PROKOP a Stavros STAVRAKIS. Exploring mechanism of enzyme catalysis by on-chip transient kinetics coupled with global data analysis and molecular modeling. \textit{Chem}. Cambridge: Cell Press, 2021, roč.~7, č.~4, s.~1066-1079. ISSN~2451-9294. Dostupné z: https://doi.org/10.1016/j.chempr.2021.02.011.

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