S-(4-bromo-2,3-dioxobutyl)glutathione: A new affinity label for the 4-4 isoenzyme of rat liver glutathione S-transferase
- Univ. of Delaware, Newark (United States)
S-(4-Bromo-2,3-dioxobutyl)glutathione (S-BDB-G), a reactive analogue of glutathione, has been synthesized and characterized by UV spectroscopy and thin-layer chromatography, as well as by bromide and primary amine analysis. Incubation of S-BDB-G (200 {mu}M) with the 4-4 isoenzyme of rat liver glutathione S-transferase at pH 6.5 and 25C results in a time-dependent inactivation of the enzyme. The k{sub obs} exhibits a nonlinear dependence on S-BDB-G concentration from 50 to 1000 {mu}M. Modified enzyme, prepared by incubating glutathione S-transferase with ({sup 3}H)S-BDB-G in the absence or in the presence of S-hexylglutathione, was reduced with NaBH{sub 4}, carboxymethylated, and digested with trypsin. The tryptic digest was fractionated by reverse-phase high-performance liquid chromatography. Two radioactive peptides were identified. These results suggest that S-BDB-G functions as an affinity label at or near the active site of glutathione S-transferase and that modification of one site per enzyme subunit causes inactivation. It is proposed that the new compound, S-(4-bromo-2,3-dioxobutyl)glutathione, may have general applicability as an affinity label of other enzymes with glutathione binding sites.
- OSTI ID:
- 5617613
- Journal Information:
- Biochemistry; (United States), Vol. 30:47; ISSN 0006-2960
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
ISOENZYMES
BIOCHEMICAL REACTION KINETICS
TRANSFERASES
INACTIVATION
ABSORPTION SPECTRA
GLUTATHIONE
LIVER
RATS
THIN-LAYER CHROMATOGRAPHY
TRITIUM COMPOUNDS
ANIMALS
BODY
CHROMATOGRAPHY
DIGESTIVE SYSTEM
DRUGS
ENZYMES
GLANDS
HYDROGEN COMPOUNDS
KINETICS
MAMMALS
ORGANIC COMPOUNDS
ORGANS
PEPTIDES
POLYPEPTIDES
PROTEINS
RADIOPROTECTIVE SUBSTANCES
REACTION KINETICS
RODENTS
SEPARATION PROCESSES
SPECTRA
VERTEBRATES
550201* - Biochemistry- Tracer Techniques