Biochemical characterization of broad-specificity enzymes using multivariate experimental design and a colorimetric microplate assay: characterization of the haloalkane dehalogenase mutants
Language English Country Netherlands Media print
Document type Evaluation Study, Journal Article, Research Support, Non-U.S. Gov't
PubMed
11165344
DOI
10.1016/s0167-7012(00)00250-5
PII: S0167701200002505
Knihovny.cz E-resources
- MeSH
- Hydrolases genetics metabolism MeSH
- Colorimetry methods MeSH
- Hydrogen-Ion Concentration MeSH
- Multivariate Analysis MeSH
- Mutation * MeSH
- Protein Engineering methods MeSH
- Sphingomonas enzymology genetics MeSH
- Substrate Specificity MeSH
- Publication type
- Journal Article MeSH
- Evaluation Study MeSH
- Research Support, Non-U.S. Gov't MeSH
- Names of Substances
- haloalkane dehalogenase MeSH Browser
- Hydrolases MeSH
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.
References provided by Crossref.org
An Ultrasensitive Fluorescence Assay for the Detection of Halides and Enzymatic Dehalogenation
Cloning, biochemical properties, and distribution of mycobacterial haloalkane dehalogenases
