Expansion of Access Tunnels and Active-Site Cavities Influence
Activity of Haloalkane Dehalogenases in Organic Cosolvents.

J 2013

Expansion of Access Tunnels and Active-Site Cavities Influence Activity of Haloalkane Dehalogenases in Organic Cosolvents.

ŠTĚPÁNKOVÁ, Veronika, M. KHABIRI, Jan BREZOVSKÝ, Antonín PAVELKA, J. SYKORA et. al.

Základní údaje

Originální název

Expansion of Access Tunnels and Active-Site Cavities Influence Activity of Haloalkane Dehalogenases in Organic Cosolvents.

Autoři

ŠTĚPÁNKOVÁ, Veronika (203 Česká republika, domácí), M. KHABIRI (364 Írán), Jan BREZOVSKÝ (203 Česká republika, domácí), Antonín PAVELKA (203 Česká republika, domácí), J. SYKORA (203 Česká republika), M. AMARO (364 Írán), B. MINOFAR (364 Írán), Zbyněk PROKOP (203 Česká republika, domácí), M. HOF (203 Česká republika), R. ETTRICH (276 Německo), Radka CHALOUPKOVÁ (203 Česká republika, domácí) a Jiří DAMBORSKÝ (203 Česká republika, garant, domácí)

Vydání

ChemBioChem, WEINHEIM, WILEY-VCH, 2013, 1439-4227

Další údaje

Jazyk

angličtina

Typ výsledku

Článek v odborném periodiku

Obor

10600 1.6 Biological sciences

Stát vydavatele

Německo

Utajení

není předmětem státního či obchodního tajemství

Impakt faktor

Impact factor: 3.060

Kód RIV

RIV/00216224:14310/13:00065821

Organizační jednotka

Přírodovědecká fakulta

DOI

http://dx.doi.org/10.1002/cbic.201200733

UT WoS

000318280500015

Klíčová slova anglicky

haloalkane dehalogenases

Štítky

AKR, rivok

Příznaky

Mezinárodní význam, Recenzováno

Změněno: 29. 4. 2014 14:21, Ing. Zdeňka Rašková

Anotace

V originále

The use of enzymes for biocatalysis can be significantly enhanced by using organic cosolvents in the reaction mixtures. Selection of the cosolvent type and concentration range for an enzymatic reaction is challenging and requires extensive empirical testing. An understanding of protein-solvent interaction could provide a theoretical framework for rationalising the selection process. Here, the behaviour of three model enzymes (haloalkane dehalogenases) was investigated in the presence of three representative organic cosolvents (acetone, formamide, and isopropanol). Steady-state kinetics assays, molecular dynamics simulations, and time-resolved fluorescence spectroscopy were used to elucidate the molecular mechanisms of enzyme-solvent interactions. Cosolvent molecules entered the enzymes’ access tunnels and active sites, enlarged their volumes with no change in overall protein structure, but surprisingly did not act as competitive inhibitors. At low concentrations, the cosolvents either enhanced catalysis by lowering K0.5 and increasing kcat , or caused enzyme inactivation by promoting substrate inhibition and decreasing kcat . The induced activation and inhibition of the enzymes correlated with expansion of the active-site pockets and their occupancy by cosolvent molecules. The study demonstrates that quantitative analysis of the proportions of the access tunnels and active-sites occupied by organic solvent molecules provides the valuable information for rational selection of appropriate protein-solvent pair and effective cosolvent concentration.

Návaznosti

ED0001/01/01, projekt VaV

Název: CETOCOEN

GAP503/12/0572, projekt VaV

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Investor: Grantová agentura ČR, Konstrukce syntetické metabolické dráhy pro degradaci důležitého environmentlního polutantu proteinovým a metabolickým inženýrstvím

GA203/08/0114, projekt VaV

Název: Specifické iontové efekty pro proteiny v roztocích a podobné biologicky relevantní systémy.

Investor: Grantová agentura ČR, Specifické iontové efekty pro proteiny v roztocích a podobné biologicky relevantní systémy

IAA401630901, projekt VaV

Název: Evoluce substrátové specifity u enzymů aktivních s xenobiotickými látkami

Investor: Akademie věd ČR, Evoluce substrátové specifity u enzymů aktivních s xenobiotickými látkami

Citovat

ŠTĚPÁNKOVÁ, Veronika, M. KHABIRI, Jan BREZOVSKÝ, Antonín PAVELKA, J. SYKORA, M. AMARO, B. MINOFAR, Zbyněk PROKOP, M. HOF, R. ETTRICH, Radka CHALOUPKOVÁ a Jiří DAMBORSKÝ. Expansion of Access Tunnels and Active-Site Cavities Influence Activity of Haloalkane Dehalogenases in Organic Cosolvents. ChemBioChem. WEINHEIM: WILEY-VCH, 2013, roč. 14, č. 7, s. 890-897. ISSN 1439-4227. Dostupné z: https://dx.doi.org/10.1002/cbic.201200733.

@article{1138864,
   author = {Štěpánková, Veronika and Khabiri, M. and Brezovský, Jan and Pavelka, Antonín and Sykora, J. and Amaro, M. and Minofar, B. and Prokop, Zbyněk and Hof, M. and Ettrich, R. and Chaloupková, Radka and Damborský, Jiří},
   article_location = {WEINHEIM},
   article_number = {7},
   doi = {http://dx.doi.org/10.1002/cbic.201200733},
   keywords = {haloalkane dehalogenases},
   language = {eng},
   issn = {1439-4227},
   journal = {ChemBioChem},
   title = {Expansion of Access Tunnels and Active-Site Cavities Influence Activity of Haloalkane Dehalogenases in Organic Cosolvents.},
   volume = {14},
   year = {2013}
}

TY  - JOUR
ID  - 1138864
AU  - Štěpánková, Veronika - Khabiri, M. - Brezovský, Jan - Pavelka, Antonín - Sykora, J. - Amaro, M. - Minofar, B. - Prokop, Zbyněk - Hof, M. - Ettrich, R. - Chaloupková, Radka - Damborský, Jiří
PY  - 2013
TI  - Expansion of Access Tunnels and Active-Site Cavities Influence Activity of Haloalkane Dehalogenases in Organic Cosolvents.
JF  - ChemBioChem
VL  - 14
IS  - 7
SP  - 890-897
EP  - 890-897
PB  - WILEY-VCH
SN  - 14394227
KW  - haloalkane dehalogenases
N2  - The use of enzymes for biocatalysis can be significantly enhanced by using organic cosolvents in the reaction mixtures. Selection of the cosolvent type and concentration range for an enzymatic reaction is challenging and requires extensive empirical testing. An understanding of protein-solvent interaction could provide a theoretical framework for rationalising the selection process. Here, the behaviour of three model enzymes (haloalkane dehalogenases) was investigated in the presence of three representative organic cosolvents (acetone, formamide, and isopropanol). Steady-state kinetics assays, molecular dynamics simulations, and time-resolved fluorescence spectroscopy were used to elucidate the molecular mechanisms of enzyme-solvent interactions. Cosolvent molecules entered the enzymes’ access tunnels and active sites, enlarged their volumes with no change in overall protein structure, but surprisingly did not act as competitive inhibitors. At low concentrations, the cosolvents either enhanced catalysis by lowering K0.5 and increasing kcat , or caused enzyme inactivation by promoting substrate inhibition and decreasing kcat . The induced activation and inhibition of the enzymes correlated with expansion of the active-site pockets and their occupancy by cosolvent molecules. The study demonstrates that quantitative analysis of the proportions of the access tunnels and active-sites occupied by organic solvent molecules provides the valuable information for rational selection of appropriate protein-solvent pair and effective cosolvent concentration.
ER  -

ŠTĚPÁNKOVÁ, Veronika, M. KHABIRI, Jan BREZOVSKÝ, Antonín PAVELKA, J. SYKORA, M. AMARO, B. MINOFAR, Zbyněk PROKOP, M. HOF, R. ETTRICH, Radka CHALOUPKOVÁ a Jiří DAMBORSKÝ. Expansion of Access Tunnels and Active-Site Cavities Influence Activity of Haloalkane Dehalogenases in Organic Cosolvents. \textit{ChemBioChem}. WEINHEIM: WILEY-VCH, 2013, roč.~14, č.~7, s.~890-897. ISSN~1439-4227. Dostupné z: https://dx.doi.org/10.1002/cbic.201200733.

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