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High-performance liquid chromatography-based assay for glutathione transferase theta 2 activity: Application to characterize interindividual variability in human liver fractions.
- Source :
-
Journal of pharmaceutical and biomedical analysis [J Pharm Biomed Anal] 2018 Jul 15; Vol. 156, pp. 181-188. Date of Electronic Publication: 2018 Apr 23. - Publication Year :
- 2018
-
Abstract
- Human glutathione transferase T2-2 (GSTT2-2) is one of the enzymes considered to play a role in inactivation of toxicants and carcinogens. The expression level of this enzyme is determined by genetic and environmental factors, which may lead to differences in susceptibility. As a specific assay for GSTT2-2 so far a spectroscopical assay based on GSH-conjugation of menaphthyl sulfate (MSu) was used. This spectrophotometric assay, however, appeared too insensitive to accurately quantify the GSTT2-2 activities in a panel of 20 human liver samples. More recently, expression levels of GSTT2-2 in biological samples are quantified by measuring mRNA levels. Since mRNA-levels do not always correlate well with enzyme activity, a specific and sensitive assay is required. In the present study a highly sensitive high-performance liquid chromatography (HPLC)-based method was developed. By applying the new method, firstly, the specificity of GSTT2-2 among 15 recombinant human GST isoforms in catalyzing GSH-conjugation of MSu was confirmed. In addition, a 65-fold inter-individual variation of GSTT2-2 activity was found from the individual liver fractions. By applying the method to individual liver fractions, a 65-fold inter-individual variation of GSTT2-2 activity was found. As a second application, the role of GSTT2-2 in GSH-conjugation of the environmental carcinogen 1-methylpyrene sulfate (MPS) was studied by correlation analysis with GSTT2-2-catalyzed MSu conjugation. The relatively poor correlation suggested that other GSTs also contribute to MPS-conjugation, as confirmed by incubations with recombinant GSTs.<br /> (Copyright © 2018 Elsevier B.V. All rights reserved.)
- Subjects :
- Biological Variation, Population
Carcinogens, Environmental metabolism
Chromatography, High Pressure Liquid methods
Glutathione metabolism
Glutathione Transferase chemistry
Glutathione Transferase metabolism
Humans
Pyrenes metabolism
Recombinant Proteins analysis
Recombinant Proteins chemistry
Recombinant Proteins metabolism
Substrate Specificity
Enzyme Assays methods
Glutathione Transferase analysis
Liver enzymology
Subjects
Details
- Language :
- English
- ISSN :
- 1873-264X
- Volume :
- 156
- Database :
- MEDLINE
- Journal :
- Journal of pharmaceutical and biomedical analysis
- Publication Type :
- Academic Journal
- Accession number :
- 29709785
- Full Text :
- https://doi.org/10.1016/j.jpba.2018.04.037