The Effect of a Unique Halide-Stabilising Residue on the
Catalytic Properties of Haloalkane Dehalogenase DatA from
Agrobacterium tumefaciens C58

J 2013

The Effect of a Unique Halide-Stabilising Residue on the Catalytic Properties of Haloalkane Dehalogenase DatA from Agrobacterium tumefaciens C58

HASAN, Khomaini, Artur Wiktor GORA, Jan BREZOVSKÝ, Radka CHALOUPKOVÁ, Hana MOSKALÍKOVÁ et. al.

Základní údaje

Originální název

The Effect of a Unique Halide-Stabilising Residue on the Catalytic Properties of Haloalkane Dehalogenase DatA from Agrobacterium tumefaciens C58

Autoři

HASAN, Khomaini (360 Indonésie, domácí), Artur Wiktor GORA (616 Polsko, domácí), Jan BREZOVSKÝ (203 Česká republika, domácí), Radka CHALOUPKOVÁ (203 Česká republika, domácí), Hana MOSKALÍKOVÁ (203 Česká republika, domácí), Andrea FOŘTOVÁ (203 Česká republika, domácí), Yuji NAGATA (392 Japonsko), Jiří DAMBORSKÝ (203 Česká republika, garant, domácí) a Zbyněk PROKOP (203 Česká republika, domácí)

Vydání

FEBS Journal, Blackwell, 2013, 1742-464X

Další údaje

Jazyk

angličtina

Typ výsledku

Článek v odborném periodiku

Obor

10600 1.6 Biological sciences

Stát vydavatele

Spojené státy

Utajení

není předmětem státního či obchodního tajemství

Impakt faktor

Impact factor: 3.986

Kód RIV

RIV/00216224:14310/13:00065822

Organizační jednotka

Přírodovědecká fakulta

DOI

http://dx.doi.org/10.1111/febs.12238

UT WoS

000320557100016

Klíčová slova anglicky

Haloalkane Dehalogenase

Štítky

AKR, rivok

Změněno: 29. 4. 2014 14:23, Ing. Zdeňka Rašková

Anotace

V originále

Haloalkane dehalogenases catalyse the hydrolysis of carbon-halogen bonds in various chlorinated, brominated and iodinated compounds. These enzymes have a conserved pair of halide-stabilising residues that are important in substrate binding and stabilisation of the transition state and the halide ion product via hydrogen bonding. In all previously known haloalkane dehalogenase, these residues are either a pair of tryptophans or a tryptophan-asparagine pair. The newly isolated haloalkane dehalogenase DatA from Agrobacterium tumefaciens C58 possesses a unique halide-stabilising tyrosine residue, Y109, in place of the conventional tryptophan. A variant of DatA with the Y109W mutation was created and the effects of this mutation on the enzyme’s structure and catalytic properties were studied using spectroscopy and pre-steady-state kinetic experiments. Quantum mechanical and molecular dynamics calculations were used to obtain a detailed analysis of the hydrogen bonding patterns within the active sites of the wild-type and the mutant, and of the stabilisation of the ligands as the reaction proceeds. Fluorescence quenching experiments suggested that replacing the tyrosine with tryptophan improves halide binding 3.7-fold, presumably due to the introduction of an additional hydrogen bond. Kinetic analysis revealed that the mutation affected the enzyme’s substrate specificity and reduced its K0.5 for selected halogenated substrates by a factor of 2-4, without impacting the rate-determining hydrolytic step. We conclude that DatA is the first natural haloalkane dehalogenase that stabilises its substrate in the active site using only a single hydrogen bond, which is a new paradigm in catalysis by this enzyme family.

Návaznosti

ED0001/01/01, projekt VaV

Název: CETOCOEN

GAP207/12/0775, projekt VaV

Název: Strukturně-funkční vztahy haloalkan dehalogenas

Investor: Grantová agentura ČR, Structure-functional Relationships of Haloalkane Dehalogenases

GAP503/12/0572, projekt VaV

Název: Konstrukce syntetické metabolické dráhy pro degradaci důležitého environmentálního polutantu proteinovým a metabolickým inženýrstvím

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

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

HASAN, Khomaini, Artur Wiktor GORA, Jan BREZOVSKÝ, Radka CHALOUPKOVÁ, Hana MOSKALÍKOVÁ, Andrea FOŘTOVÁ, Yuji NAGATA, Jiří DAMBORSKÝ a Zbyněk PROKOP. The Effect of a Unique Halide-Stabilising Residue on the Catalytic Properties of Haloalkane Dehalogenase DatA from Agrobacterium tumefaciens C58. FEBS Journal. Blackwell, 2013, roč. 280, č. 13, s. 3149-3159. ISSN 1742-464X. Dostupné z: https://dx.doi.org/10.1111/febs.12238.

@article{1138865,
   author = {Hasan, Khomaini and Gora, Artur Wiktor and Brezovský, Jan and Chaloupková, Radka and Moskalíková, Hana and Fořtová, Andrea and Nagata, Yuji and Damborský, Jiří and Prokop, Zbyněk},
   article_number = {13},
   doi = {http://dx.doi.org/10.1111/febs.12238},
   keywords = {Haloalkane Dehalogenase},
   language = {eng},
   issn = {1742-464X},
   journal = {FEBS Journal},
   title = {The Effect of a Unique Halide-Stabilising Residue on the Catalytic Properties of Haloalkane Dehalogenase DatA from Agrobacterium tumefaciens C58},
   volume = {280},
   year = {2013}
}

TY  - JOUR
ID  - 1138865
AU  - Hasan, Khomaini - Gora, Artur Wiktor - Brezovský, Jan - Chaloupková, Radka - Moskalíková, Hana - Fořtová, Andrea - Nagata, Yuji - Damborský, Jiří - Prokop, Zbyněk
PY  - 2013
TI  - The Effect of a Unique Halide-Stabilising Residue on the Catalytic Properties of Haloalkane Dehalogenase DatA from Agrobacterium tumefaciens C58
JF  - FEBS Journal
VL  - 280
IS  - 13
SP  - 3149-3159
EP  - 3149-3159
PB  - Blackwell
SN  - 1742464X
KW  - Haloalkane Dehalogenase
N2  - Haloalkane dehalogenases catalyse the hydrolysis of carbon-halogen bonds in various chlorinated, brominated and iodinated compounds. These enzymes have a conserved pair of halide-stabilising residues that are important in substrate binding and stabilisation of the transition state and the halide ion product via hydrogen bonding. In all previously known haloalkane dehalogenase, these residues are either a pair of tryptophans or a tryptophan-asparagine pair. The newly isolated haloalkane dehalogenase DatA from Agrobacterium tumefaciens C58 possesses a unique halide-stabilising tyrosine residue, Y109, in place of the conventional tryptophan. A variant of DatA with the Y109W mutation was created and the effects of this mutation on the enzyme’s structure and catalytic properties were studied using spectroscopy and pre-steady-state kinetic experiments. Quantum mechanical and molecular dynamics calculations were used to obtain a detailed analysis of the hydrogen bonding patterns within the active sites of the wild-type and the mutant, and of the stabilisation of the ligands as the reaction proceeds. Fluorescence quenching experiments suggested that replacing the tyrosine with tryptophan improves halide binding 3.7-fold, presumably due to the introduction of an additional hydrogen bond. Kinetic analysis revealed that the mutation affected the enzyme’s substrate specificity and reduced its K0.5 for selected halogenated substrates by a factor of 2-4, without impacting the rate-determining hydrolytic step. We conclude that DatA is the first natural haloalkane dehalogenase that stabilises its substrate in the active site using only a single hydrogen bond, which is a new paradigm in catalysis by this enzyme family.
ER  -

HASAN, Khomaini, Artur Wiktor GORA, Jan BREZOVSKÝ, Radka CHALOUPKOVÁ, Hana MOSKALÍKOVÁ, Andrea FOŘTOVÁ, Yuji NAGATA, Jiří DAMBORSKÝ a Zbyněk PROKOP. The Effect of a Unique Halide-Stabilising Residue on the Catalytic Properties of Haloalkane Dehalogenase DatA from Agrobacterium tumefaciens C58. \textit{FEBS Journal}. Blackwell, 2013, roč.~280, č.~13, s.~3149-3159. ISSN~1742-464X. Dostupné z: https://dx.doi.org/10.1111/febs.12238.

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