Exploring the challenges of computational enzyme design by
rebuilding the active site of a dehalogenase

J 2019

Exploring the challenges of computational enzyme design by rebuilding the active site of a dehalogenase

JINDAL, Garima; Kateřina SLÁNSKÁ; Veselin KOLEV; Jiří DAMBORSKÝ; Zbyněk PROKOP et. al.

Základní údaje

Originální název

Exploring the challenges of computational enzyme design by rebuilding the active site of a dehalogenase

Autoři

JINDAL, Garima (356 Indie); Kateřina SLÁNSKÁ (203 Česká republika, domácí); Veselin KOLEV (840 Spojené státy); Jiří DAMBORSKÝ (203 Česká republika, garant, domácí); Zbyněk PROKOP (203 Česká republika, domácí) a Arieh WARSHEL (840 Spojené státy)

Vydání

Proceedings of the National Academy of Sciences of the United States of America, WASHINGTON, NATL ACAD SCIENCES, 2019, 0027-8424

Další údaje

Jazyk

angličtina

Typ výsledku

Článek v odborném periodiku

Obor

10608 Biochemistry and molecular biology

Stát vydavatele

Spojené státy

Utajení

není předmětem státního či obchodního tajemství

Odkazy

URL

Impakt faktor

Impact factor: 9.412

Kód RIV

RIV/00216224:14310/19:00108175

Organizační jednotka

Přírodovědecká fakulta

DOI

http://dx.doi.org/10.1073/pnas.1804979115

UT WoS

000455086900011

EID Scopus

2-s2.0-85059652342

Klíčová slova anglicky

enzyme design; EVB; transient kinetics; dehalogenase; nucleophilic substitution

Štítky

rivok

Příznaky

Mezinárodní význam, Recenzováno

Změněno: 16. 2. 2023 12:12, Mgr. Michaela Hylsová, Ph.D.

Anotace

V originále

Rational enzyme design presents a major challenge that has not been overcome by computational approaches. One of the key challenges is the difficulty in assessing the magnitude of the maximum possible catalytic activity. In an attempt to overcome this challenge, we introduce a strategy that takes an active enzyme (assuming that its activity is close to the maximum possible activity), design mutations that reduce the catalytic activity, and then try to restore that catalysis by mutating other residues. Here we take as a test case the enzyme haloalkane dehalogenase (DhlA), with a 1,2-dichloroethane substrate. We start by demonstrating our ability to reproduce the results of single mutations. Next, we design mutations that reduce the enzyme activity and finally design double mutations that are aimed at restoring the activity. Using the computational predictions as a guide, we conduct an experimental study that confirms our prediction in one case and leads to inconclusive results in another case with 1,2-dichloroethane as substrate. Interestingly, one of our predicted double mutants catalyzes dehalogenation of 1,2-dibromoethane more efficiently than the wild-type enzyme.

Návaznosti

GA16-07965S, projekt VaV

Název: Řízená evoluce dynamických elementů v enzymech s využitím mikrofluidních čipů

Investor: Grantová agentura ČR, Řízená evoluce dynamických elementů v enzymech s využitím mikrofluidních čipů

LM2015047, projekt VaV

Název: Česká národní infrastruktura pro biologická data (Akronym: ELIXIR-CZ)

Investor: Ministerstvo školství, mládeže a tělovýchovy ČR, Česká národní infrastruktura pro biologická data

LM2015051, projekt VaV

Název: Centrum pro výzkum toxických látek v prostředí (Akronym: RECETOX RI)

Investor: Ministerstvo školství, mládeže a tělovýchovy ČR, Výzkumná infrastruktura RECETOX

LM2015055, projekt VaV

Název: Centrum pro systémovou biologii (Akronym: C4SYS)

Investor: Ministerstvo školství, mládeže a tělovýchovy ČR, The national infrastructure C4SYS - Centre for Systems Biology

LO1214, projekt VaV

Název: Centrum pro výzkum toxických látek v prostředí (Akronym: RECETOX)

Investor: Ministerstvo školství, mládeže a tělovýchovy ČR, Centrum pro výzkum toxických látek v prostředí

Citovat

JINDAL, Garima; Kateřina SLÁNSKÁ; Veselin KOLEV; Jiří DAMBORSKÝ; Zbyněk PROKOP a Arieh WARSHEL. Exploring the challenges of computational enzyme design by rebuilding the active site of a dehalogenase. Proceedings of the National Academy of Sciences of the United States of America. WASHINGTON: NATL ACAD SCIENCES, 2019, roč. 116, č. 2, s. 389-394. ISSN 0027-8424. Dostupné z: https://dx.doi.org/10.1073/pnas.1804979115.

@article{1632620,
   author = {Jindal, Garima and Slánská, Kateřina and Kolev, Veselin and Damborský, Jiří and Prokop, Zbyněk and Warshel, Arieh},
   article_location = {WASHINGTON},
   article_number = {2},
   doi = {http://dx.doi.org/10.1073/pnas.1804979115},
   keywords = {enzyme design; EVB; transient kinetics; dehalogenase; nucleophilic substitution},
   language = {eng},
   issn = {0027-8424},
   journal = {Proceedings of the National Academy of Sciences of the United States of America},
   title = {Exploring the challenges of computational enzyme design by rebuilding the active site of a dehalogenase},
   url = {http://dx.doi.org/10.1073/pnas.1804979115},
   volume = {116},
   year = {2019}
}

TY  - JOUR
ID  - 1632620
AU  - Jindal, Garima - Slánská, Kateřina - Kolev, Veselin - Damborský, Jiří - Prokop, Zbyněk - Warshel, Arieh
PY  - 2019
TI  - Exploring the challenges of computational enzyme design by rebuilding the active site of a dehalogenase
JF  - Proceedings of the National Academy of Sciences of the United States of America
VL  - 116
IS  - 2
SP  - 389-394
EP  - 389-394
PB  - NATL ACAD SCIENCES
SN  - 00278424
KW  - enzyme design
KW  - EVB
KW  - transient kinetics
KW  - dehalogenase
KW  - nucleophilic substitution
UR  - http://dx.doi.org/10.1073/pnas.1804979115
L2  - http://dx.doi.org/10.1073/pnas.1804979115
N2  - Rational enzyme design presents a major challenge that has not been overcome by computational approaches. One of the key challenges is the difficulty in assessing the magnitude of the maximum possible catalytic activity. In an attempt to overcome this challenge, we introduce a strategy that takes an active enzyme (assuming that its activity is close to the maximum possible activity), design mutations that reduce the catalytic activity, and then try to restore that catalysis by mutating other residues. Here we take as a test case the enzyme haloalkane dehalogenase (DhlA), with a 1,2-dichloroethane substrate. We start by demonstrating our ability to reproduce the results of single mutations. Next, we design mutations that reduce the enzyme activity and finally design double mutations that are aimed at restoring the activity. Using the computational predictions as a guide, we conduct an experimental study that confirms our prediction in one case and leads to inconclusive results in another case with 1,2-dichloroethane as substrate. Interestingly, one of our predicted double mutants catalyzes dehalogenation of 1,2-dibromoethane more efficiently than the wild-type enzyme.
ER  -

JINDAL, Garima; Kateřina SLÁNSKÁ; Veselin KOLEV; Jiří DAMBORSKÝ; Zbyněk PROKOP a Arieh WARSHEL. Exploring the challenges of computational enzyme design by rebuilding the active site of a dehalogenase. \textit{Proceedings of the National Academy of Sciences of the United States of America}. WASHINGTON: NATL ACAD SCIENCES, 2019, roč.~116, č.~2, s.~389-394. ISSN~0027-8424. Dostupné z: https://dx.doi.org/10.1073/pnas.1804979115.

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