Biochemical characterization of broad-specificity enzymes using multivariate experimental design and a colorimetric microplate assay: characterization of the haloalkane dehalogenase mutants

MARVANOVÁ, Soňa, Yuji NAGATA, Michaela WIMMEROVÁ, Jana SÝKOROVÁ, Kamila HYNKOVÁ and Jiří DAMBORSKÝ. Biochemical characterization of broad-specificity enzymes using multivariate experimental design and a colorimetric microplate assay: characterization of the haloalkane dehalogenase mutants. Journal of Microbiological Methods. 2001, vol. 44, No 1, p. 149-157. ISSN 0167-7012.

Basic information

• Original name: Biochemical characterization of broad-specificity enzymes using multivariate experimental design and a colorimetric microplate assay: characterization of the haloalkane dehalogenase mutants
• Authors: MARVANOVÁ, Soňa, Yuji NAGATA, Michaela WIMMEROVÁ, Jana SÝKOROVÁ, Kamila HYNKOVÁ and Jiří DAMBORSKÝ.
• Edition: Journal of Microbiological Methods, 2001, 0167-7012.

Other information

• Original language: English
• Type of outcome: Article in a journal
• Field of Study: 10600 1.6 Biological sciences
• Country of publisher: Netherlands
• Confidentiality degree: is not subject to a state or trade secret
• WWW: URL
• Impact factor: Impact factor: 1.810
• RIV identification code: RIV/00216224:14310/01:00002964
• Organization unit: Faculty of Science
• UT WoS: 000167080200007

Abstract

The pH indicator dye-based colorimetric method and multivariate experimental design were used for the systematic biochemical characterization of the broad-specificity enzymes haloalkane dehalogenases. Halogenated compounds for characterization of the enzymes were selected using Principal Component Analysis. The substrates were characterised by 24 physico-chemical and structural descriptors. Thirty four substrates were selected for testing out of 194 halogenated compounds. Relative activities determined using the optimised colorimetric microplate assay were validated against the catalytic constants determined by gas chromatography. The applicability of the assay was tested with F151L, F154L and F169L mutants of the haloalkane dehalogenase from Sphingomonas paucimobilis UT26.

Links

• ME 276, research and development project: Name: Racionální re-design mikrobiálních enzymů podílejících se na degradaci toxických organických polutantů

Investor: Ministry of Education, Youth and Sports of the CR, Rational re-design of microbial enzymes involved in degradation of toxic organic pollutants.

• MSM 143100005, plan (intention): Name: Strukturně-funkční vztahy biomolekul a jejich role v metabolismu

Investor: Ministry of Education, Youth and Sports of the CR, Biomolecular Structure-function Relationships and their role in the Metabolism