Mechanism-guided tunnel engineering to increase the efficiency
of a flavin-dependent halogenase

J 2022

Mechanism-guided tunnel engineering to increase the efficiency of a flavin-dependent halogenase

PRAKINEE, Kridsadakorn, Aisaraphon PHINTHA, Surawit VISITSATTHAWONG, Narin LAWAN, Jeerus SUCHARITAKUL et. al.

Základní údaje

Originální název

Mechanism-guided tunnel engineering to increase the efficiency of a flavin-dependent halogenase

Autoři

PRAKINEE, Kridsadakorn, Aisaraphon PHINTHA, Surawit VISITSATTHAWONG, Narin LAWAN, Jeerus SUCHARITAKUL, Chadaporn KANTIWIRIYAWANITCH, Jiří DAMBORSKÝ (203 Česká republika, garant, domácí), Penchit CHITNUMSUB, Van Pee KARL-HEINZ a Pimchai CHAIYEN

Vydání

NATURE CATALYSIS, ENGLAND, NATURE PORTFOLIO, 2022, 2520-1158

Další údaje

Jazyk

angličtina

Typ výsledku

Článek v odborném periodiku

Obor

10403 Physical chemistry

Stát vydavatele

Německo

Utajení

není předmětem státního či obchodního tajemství

Odkazy

URL

Impakt faktor

Impact factor: 37.800

Kód RIV

RIV/00216224:14310/22:00126374

Organizační jednotka

Přírodovědecká fakulta

DOI

http://dx.doi.org/10.1038/s41929-022-00800-8

UT WoS

000812063200001

Klíčová slova anglicky

TRYPTOPHAN 7-HALOGENASE; BIOCATALYTIC SCOPE; MOLECULAR-DYNAMICS; KINETIC MECHANISM; CHLORINATION; BIOSYNTHESIS; INTERMEDIATE; INSIGHTS; PHENOL; REBH

Štítky

rivok

Příznaky

Mezinárodní význam, Recenzováno

Změněno: 16. 3. 2023 21:52, Mgr. Michaela Hylsová, Ph.D.

Anotace

V originále

Although flavin-dependent halogenases (FDHs) are attractive for C-H bond activation, their applications are limited due to low turnover and stability. We have previously shown that leakage of a halogenating intermediate, hypohalous acid (HOX), causes FDHs to be inefficient by lessening halogenation yield. Here we employed a mechanism-guided semi-rational approach to engineer the intermediate transfer tunnel connecting two active sites of tryptophan 6-halogenase (Thal). This Thal-V82I variant generates less HOX leakage and possesses multiple catalytic improvements such as faster halogenation, broader substrate utilization, and greater thermostability and pH tolerance compared with the wildtype Thal. Stopped-flow and rapid quench kinetics analyses indicated that rate constants of halogenation and flavin oxidation are faster for Thal-V82I. Molecular dynamics simulations revealed that the V82I substitution introduces hydrophobic interactions which regulate tunnel dynamics to accommodate HOX and cause rearrangement of water networks, allowing better use of various substrates than the wildtype. Our approach demonstrates that an in-depth understanding of reaction mechanisms is valuable for improving efficiency of FDHs.

Návaznosti

LM2018121, projekt VaV

Název: Výzkumná infrastruktura RECETOX (Akronym: RECETOX RI)

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

Citovat

PRAKINEE, Kridsadakorn, Aisaraphon PHINTHA, Surawit VISITSATTHAWONG, Narin LAWAN, Jeerus SUCHARITAKUL, Chadaporn KANTIWIRIYAWANITCH, Jiří DAMBORSKÝ, Penchit CHITNUMSUB, Van Pee KARL-HEINZ a Pimchai CHAIYEN. Mechanism-guided tunnel engineering to increase the efficiency of a flavin-dependent halogenase. NATURE CATALYSIS. ENGLAND: NATURE PORTFOLIO, 2022, roč. 5, č. 6, s. 534-544. ISSN 2520-1158. Dostupné z: https://dx.doi.org/10.1038/s41929-022-00800-8.

@article{1883608,
   author = {Prakinee, Kridsadakorn and Phintha, Aisaraphon and Visitsatthawong, Surawit and Lawan, Narin and Sucharitakul, Jeerus and Kantiwiriyawanitch, Chadaporn and Damborský, Jiří and Chitnumsub, Penchit and KarlandHeinz, Van Pee and Chaiyen, Pimchai},
   article_location = {ENGLAND},
   article_number = {6},
   doi = {http://dx.doi.org/10.1038/s41929-022-00800-8},
   keywords = {TRYPTOPHAN 7-HALOGENASE; BIOCATALYTIC SCOPE; MOLECULAR-DYNAMICS; KINETIC MECHANISM; CHLORINATION; BIOSYNTHESIS; INTERMEDIATE; INSIGHTS; PHENOL; REBH},
   language = {eng},
   issn = {2520-1158},
   journal = {NATURE CATALYSIS},
   title = {Mechanism-guided tunnel engineering to increase the efficiency of a flavin-dependent halogenase},
   url = {https://www.nature.com/articles/s41929-022-00800-8},
   volume = {5},
   year = {2022}
}

TY  - JOUR
ID  - 1883608
AU  - Prakinee, Kridsadakorn - Phintha, Aisaraphon - Visitsatthawong, Surawit - Lawan, Narin - Sucharitakul, Jeerus - Kantiwiriyawanitch, Chadaporn - Damborský, Jiří - Chitnumsub, Penchit - Karl-Heinz, Van Pee - Chaiyen, Pimchai
PY  - 2022
TI  - Mechanism-guided tunnel engineering to increase the efficiency of a flavin-dependent halogenase
JF  - NATURE CATALYSIS
VL  - 5
IS  - 6
SP  - 534-544
EP  - 534-544
PB  - NATURE PORTFOLIO
SN  - 25201158
KW  - TRYPTOPHAN 7-HALOGENASE
KW  - BIOCATALYTIC SCOPE
KW  - MOLECULAR-DYNAMICS
KW  - KINETIC MECHANISM
KW  - CHLORINATION
KW  - BIOSYNTHESIS
KW  - INTERMEDIATE
KW  - INSIGHTS
KW  - PHENOL
KW  - REBH
UR  - https://www.nature.com/articles/s41929-022-00800-8
N2  - Although flavin-dependent halogenases (FDHs) are attractive for C-H bond activation, their applications are limited due to low turnover and stability. We have previously shown that leakage of a halogenating intermediate, hypohalous acid (HOX), causes FDHs to be inefficient by lessening halogenation yield. Here we employed a mechanism-guided semi-rational approach to engineer the intermediate transfer tunnel connecting two active sites of tryptophan 6-halogenase (Thal). This Thal-V82I variant generates less HOX leakage and possesses multiple catalytic improvements such as faster halogenation, broader substrate utilization, and greater thermostability and pH tolerance compared with the wildtype Thal. Stopped-flow and rapid quench kinetics analyses indicated that rate constants of halogenation and flavin oxidation are faster for Thal-V82I. Molecular dynamics simulations revealed that the V82I substitution introduces hydrophobic interactions which regulate tunnel dynamics to accommodate HOX and cause rearrangement of water networks, allowing better use of various substrates than the wildtype. Our approach demonstrates that an in-depth understanding of reaction mechanisms is valuable for improving efficiency of FDHs.
ER  -

PRAKINEE, Kridsadakorn, Aisaraphon PHINTHA, Surawit VISITSATTHAWONG, Narin LAWAN, Jeerus SUCHARITAKUL, Chadaporn KANTIWIRIYAWANITCH, Jiří DAMBORSKÝ, Penchit CHITNUMSUB, Van Pee KARL-HEINZ a Pimchai CHAIYEN. Mechanism-guided tunnel engineering to increase the efficiency of a flavin-dependent halogenase. \textit{NATURE CATALYSIS}. ENGLAND: NATURE PORTFOLIO, 2022, roč.~5, č.~6, s.~534-544. ISSN~2520-1158. Dostupné z: https://dx.doi.org/10.1038/s41929-022-00800-8.

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