Radiation inactivation of microsomal glutathione S-transferase

T. D. Boyer, D. A. Vessey, E. Kempner

Research output: Contribution to journalArticle

37 Scopus citations

Abstract

Radiation inactivation analysis was used to determine the target size of rat liver microsomal glutathione S-transferase both in situ and following purification. When Tris-HCl-washed microsomes were irradiated, there was a 1.5-2.0-fold increase in enzymatic activity over the first 3-6 megarads followed by a decrease in enzymatic activity. Above 48 megarads the radiation inactivation curve of the Tris-HCl-washed microsomes was described by a monoexponential function which gave a target size of 48 kDa. The enzymatic activity of the microsomal enzyme was selectively increased by treating the Tris-HCl-washed microsomes either with N-ethylmaleimide or washing the microsomes with small unilamellar vesicles made from phosphatidylcholine. The inactivation curves obtained with both types of treated microsomes were simple monoexponential decays in enzymatic activity with target sizes of 46 kDa (N-ethylmaleimide) and 44 kDa (unilamellar vesicles). The microsomal enzyme was detergent solubilized and purified. The M(r) values of the purified protein was 15,500 (sodium dodecyl sulfate-polyacrylamide gel electrophoresis). These data suggest that the functional unit of the microsomal form of glutathione S-transferase in situ is a trimer. The target size of the purified enzyme solubilized in Triton X-100 was 85 kDa, and no increase in activity was observed at the lower radiation doses. The increase in the target size of the purified enzyme could not be ascribed solely to the presence of the detergent. This result suggests that the microsomal form of this enzyme can exist as catalytically active oligomers of different sizes depending on its environment.

Original languageEnglish (US)
Pages (from-to)16963-16968
Number of pages6
JournalJournal of Biological Chemistry
Volume261
Issue number36
StatePublished - 1986

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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