About: A Single Mutation in a Tunnel to the Active Site Changes the Mechanism and Kinetics of Product Release in Haloalkane Dehalogenase LinB

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About: A Single Mutation in a Tunnel to the Active Site Changes the Mechanism and Kinetics of Product Release in Haloalkane Dehalogenase LinB Goto Sponge NotDistinct Permalink

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rdf:type	skos:Concept http://linked.opendata.cz/ontology/domain/vavai/Vysledek
Description	Many enzymes have buried active sites. The properties of the tunnels connecting the active site with bulk solvent affect ligand binding and unbinding and also the catalytic properties. Here, we investigate ligand passage in the haloalkane dehalogenase enzyme LinB and the effect of replacing leucine by a bulky tryptophan at a tunnel-lining position. Transient kinetic experiments show that the mutation significantly slows down the rate of product release. Moreover, the mechanism of bromide ion release is changed from a one-step process in the wild type enzyme to a two-step process in the mutant. The rate constant of bromide ion release corresponds to the overall steady-state turnover rate constant, suggesting that product release became the rate-limiting step of catalysis in the mutant. We explain the experimental findings by investigating the molecular details of the process computationally. Many enzymes have buried active sites. The properties of the tunnels connecting the active site with bulk solvent affect ligand binding and unbinding and also the catalytic properties. Here, we investigate ligand passage in the haloalkane dehalogenase enzyme LinB and the effect of replacing leucine by a bulky tryptophan at a tunnel-lining position. Transient kinetic experiments show that the mutation significantly slows down the rate of product release. Moreover, the mechanism of bromide ion release is changed from a one-step process in the wild type enzyme to a two-step process in the mutant. The rate constant of bromide ion release corresponds to the overall steady-state turnover rate constant, suggesting that product release became the rate-limiting step of catalysis in the mutant. We explain the experimental findings by investigating the molecular details of the process computationally. (en)
Title	A Single Mutation in a Tunnel to the Active Site Changes the Mechanism and Kinetics of Product Release in Haloalkane Dehalogenase LinB A Single Mutation in a Tunnel to the Active Site Changes the Mechanism and Kinetics of Product Release in Haloalkane Dehalogenase LinB (en)
skos:prefLabel	A Single Mutation in a Tunnel to the Active Site Changes the Mechanism and Kinetics of Product Release in Haloalkane Dehalogenase LinB A Single Mutation in a Tunnel to the Active Site Changes the Mechanism and Kinetics of Product Release in Haloalkane Dehalogenase LinB (en)
skos:notation	RIV/00216224:14310/12:00057142!RIV13-AV0-14310___
http://linked.open...avai/predkladatel	Přírodovědecká fakulta
http://linked.open...avai/riv/aktivita	P
http://linked.open...avai/riv/aktivity	P(ED0001/01/01), P(IAA401630901)
http://linked.open...iv/cisloPeriodika	34
http://linked.open...vai/riv/dodaniDat	2013
http://linked.open...aciTvurceVysledku	Prokop, Zbyněk Chovancová, Eva Damborský, Jiří Góra, Artur Wiktor
http://linked.open.../riv/druhVysledku	J - Článek v odborném periodiku
http://linked.open...iv/duvernostUdaju	S - Úplné a pravdivé údaje nepodléhající ochraně podle zvláštních právních předpisů
http://linked.open...titaPredkladatele	Masarykova univerzita / Přírodovědecká fakulta
http://linked.open...dnocenehoVysledku	120620
http://linked.open...ai/riv/idVysledku	RIV/00216224:14310/12:00057142
http://linked.open...riv/jazykVysledku	eng - angličtina
http://linked.open.../riv/klicovaSlova	HLD; haloalkane dehalogenase; MD; molecular dynamics; RAMD; random acceleration molecular dynamics; ABF; adaptive biasing force; RC; reaction coordinate; FEP; free energy perturbation; NATA; N-acetyltryptophan amide (en)
http://linked.open.../riv/klicoveSlovo	FEP ABF HLD MD N-acetyltryptophan amide NATA RAMD RC adaptive biasing force free energy perturbation random acceleration molecular dynamics reaction coordinate molecular dynamics haloalkane dehalogenase
http://linked.open...odStatuVydavatele	US - Spojené státy americké
http://linked.open...ontrolniKodProRIV	[1CC491546B0D]
http://linked.open...i/riv/nazevZdroje	The Journal of Biological Chemistry
http://linked.open...in/vavai/riv/obor	EB
http://linked.open...ichTvurcuVysledku	4 (xsd:int)
http://linked.open...cetTvurcuVysledku	7 (xsd:int)
http://linked.open...vavai/riv/projekt	CETOCOEN Evolution of substrate specificity in enzymes acting on xenobiotic compounds
http://linked.open...UplatneniVysledku	2012
http://linked.open...v/svazekPeriodika	287
http://linked.open...iv/tvurceVysledku	Biedermannová, Lada Damborský, Jiří Góra, Artur Wiktor Prokop, Zbyněk Kovács, Mihály Wade, Rebecca C. Šebestová, Eva
http://linked.open...ain/vavai/riv/wos	000308074600073
issn	0021-9258
number of pages	13 (xsd:int)
http://bibframe.org/vocab/doi	10.1074/jbc.M112.377853
http://localhost/t...ganizacniJednotka	14310
is http://linked.open...avai/riv/vysledek of	A Single Mutation in a Tunnel to the Active Site Changes the Mechanism and Kinetics of Product Release in Haloalkane Dehalogenase LinB

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