Detailed Information on Publication Record
2009
Nanosecond Time-Dependent Stokes Shift at the Tunnel Mouth of Haloalkane Dehalogenases
FOŘTOVÁ, Andrea, J. SÝKORA, Agniezska OLZYNSKA, Jan BREZOVSKÝ, Zbyněk ZDRÁHAL et. al.Basic information
Original name
Nanosecond Time-Dependent Stokes Shift at the Tunnel Mouth of Haloalkane Dehalogenases
Name in Czech
Nanosekundová časová závislost spektrálního posunu v tunelu haloalkán dehalogenáz
Authors
FOŘTOVÁ, Andrea (203 Czech Republic, belonging to the institution), J. SÝKORA (203 Czech Republic), Agniezska OLZYNSKA (616 Poland), Jan BREZOVSKÝ (203 Czech Republic, belonging to the institution), Zbyněk ZDRÁHAL (203 Czech Republic, belonging to the institution), Jiří DAMBORSKÝ (203 Czech Republic, guarantor, belonging to the institution) and Martin HOF (203 Czech Republic)
Edition
JOURNAL OF THE AMERICAN CHEMICAL SOCIETY, 2009, 0002-7863
Other information
Language
English
Type of outcome
Článek v odborném periodiku
Field of Study
10600 1.6 Biological sciences
Country of publisher
United States of America
Confidentiality degree
není předmětem státního či obchodního tajemství
References:
Impact factor
Impact factor: 8.580
RIV identification code
RIV/00216224:14310/09:00033990
Organization unit
Faculty of Science
UT WoS
000262521800036
Keywords (in Czech)
haloalkán dehalogenázy, DBjA, DhaA
Keywords in English
tunnel mouths; haloalkane dehalogenases; DbjA-H280F and DhaA-H272F;
Tags
International impact, Reviewed
Změněno: 23/3/2012 11:05, prof. Mgr. Jiří Damborský, Dr.
V originále
The tunnel mouths are evolutionally the most variable regions in the structures of haloalkane dehalogenases originating from different bacterial species suggesting their importance for adaptation of enzymes to various substrates. We decided to monitor the dynamics of this particular region by means of time resolved fluorescence spectroscopy and molecular dynamic simulations. To label the enzyme specifically, we adapted a novel procedure that utilizes a coumarin dye containing a halide-hydrocarbon linker, which serves as a substrate for enzymatic reaction. The procedure leads to a coumarin dye covalently attached and specifically located in the tunnel mouth of the enzyme. In this manner, we stained two haloalkane dehalogenase mutants, DbjA-H280F and DhaA-H272F. The measurements of time-resolved fluorescence anisotropy, acrylamide quenching and time resolved emission spectra reveal differences in the polarity, accessibility and mobility of the dye and its microenvironment for both of the mutants. The obtained experimental data are consistent with the results obtained by molecular dynamics calculations and correlate with the anatomy of the tunnel mouths, which were proposed to have a strong impact on the catalytic activity and specificity of the examined mutants. Interestingly, the kinetics of the recorded time-dependent Stokes shift is unusual slow; it occurs on the nanosecond time-scale, suggesting that the protein dynamics is extremely slowed down at the region involved in the exchange of ligands between the active site cavity and bulk solvent.
In Czech
Článek popisuje studium dynamiky vod v proteinech s využitím fluoresecnční spektroskopie a molekulové dynamiky.
Links
| LC06010, research and development project |
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| MSM0021622412, plan (intention) |
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| MSM0021622413, plan (intention) |
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| MSM0021622415, plan (intention) |
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