Detailed Information on Publication Record
2007
Study of substrate inhibition by electrophoretically mediated microanalysis in partially filled capillary.
PAPEŽOVÁ, Kateřina, Tomáš NĚMEC, Radka CHALOUPKOVÁ and Zdeněk GLATZBasic information
Original name
Study of substrate inhibition by electrophoretically mediated microanalysis in partially filled capillary.
Name in Czech
Studium substrátové inhibice metodou EMMA v kombinaci s technikou částečného plnění kapiláry
Authors
PAPEŽOVÁ, Kateřina (203 Czech Republic), Tomáš NĚMEC (203 Czech Republic), Radka CHALOUPKOVÁ (203 Czech Republic) and Zdeněk GLATZ (203 Czech Republic, guarantor)
Edition
Journal of Chromatography A, Amsterdam, Elsevier, 2007, 0021-9673
Other information
Language
English
Type of outcome
Článek v odborném periodiku
Field of Study
10600 1.6 Biological sciences
Country of publisher
Netherlands
Confidentiality degree
není předmětem státního či obchodního tajemství
Impact factor
Impact factor: 3.641
RIV identification code
RIV/00216224:14310/07:00020038
Organization unit
Faculty of Science
UT WoS
000247049200043
Keywords in English
CE; EMMA substrate; inhibition; haloalkane dehalogenase
Tags
International impact, Reviewed
Změněno: 29/6/2008 05:54, Olga Křížová
V originále
Substrate inhibition is a common phenomenon in enzyme kinetics. We report here for the first time its study by a combination of the electrophoretically mediated microanalysis (EMMA) methodology with a partial filling technique. In this set-up, the part of capillary is filled with the buffer best for the enzymatic reaction whereas, the rest of the capillary is filled with the background electrolyte optimal for separation of substrates and products. In the case of haloalkane dehalogenase, a model enzyme selected for this study, the enzymatic reaction was performed in 20 mM glycine buffer (pH 8.6) whereas 20 mM beta-alanine - hydrochloric acid buffer (pH 3.5) was used as a background electrolyte in combination with direct detection at 200 nm. The whole study was performed on poorly soluble brominated substrate - 1,2-dibromoethane. As a result it was first necessary to find the compromise between the concentrations of the enzyme and the substrate preserving both the adequate sensitivity of the assay and at the same time the attainable substrate solubility. By means of the developed EMMA methodology we were able to determine the Michaelis constant (KM) as well as the substrate inhibition constant (KSI). The value of KM and KSI obtained were 7.7 mM and 1.1 mM respectively. Observation of the substrate inhibition of haloalkane dehalogenase by 1,2-dibromoethane is in accordance with previous literature data.
In Czech
Byla vypracována technika pro studium substrátové inhibice metodou EMMA v kombinaci s technikou částečného plnění kapiláry
Links
| GA203/06/0047, research and development project |
| ||
| LC06023, research and development project |
| ||
| MSM0021622413, plan (intention) |
|