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Identification of a hepatic plasma membrane glutathione S-transferase activated by N-ethylmaleimide
M E Horbach1, H Sies, T P Akerboom
1Institut für Physiologische Chemie I, Heinrich-Heine-Universität Düsseldorf, Germany.
Abstract:
Rat liver plasma membranes exhibit membrane-bound glutathione S-transferase activity. The specific activity in isolated canalicular membranes was 83 +/- 8 mU/mg protein and 50 +/- 3 mU/mg protein in the sinusoidal membranes. Whereas microsomal and outer mitochondrial glutathione S-transferases were stimulated seven and four-fold with N-ethylmaleimide, respectively, the plasma membrane activity was activated two-fold. Western blot analysis, using an antibody against the microsomal glutathione S-transferase, shows the presence of a 17 kDa protein in canalicular and sinusoidal membrane fractions. The antibody reaction was about three-fold higher in the canalicular compared to the sinusoidal membrane fraction. These data support the conclusion that the plasma membrane glutathione S-transferase is closely related to the microsomal and outer mitochondrial membrane enzyme.
Insights
Rat liver plasma membranes possess glutathione S-transferase activity. This enzyme is closely related to microsomal and outer mitochondrial forms, as shown by N-ethylmaleimide activation and Western blot analysis.
Area of Science:
- Biochemistry
- Cell Biology
- Enzymology
Background:
- Glutathione S-transferases (GSTs) are crucial enzymes involved in detoxification.
- Membrane-bound GSTs play roles in cellular protection and transport.
- Understanding the localization and characteristics of GSTs in rat liver membranes is important.
Purpose of the Study:
- To investigate the presence and characteristics of glutathione S-transferase activity in rat liver plasma membranes.
- To compare plasma membrane-bound GST activity with that of other cellular fractions.
- To determine the relationship between plasma membrane GST and other known GST isoforms.
Main Methods:
- Isolation of rat liver plasma membrane fractions (canalicular and sinusoidal).
- Assay of glutathione S-transferase specific activity.
- Enzyme activation studies using N-ethylmaleimide.
- Western blot analysis using an antibody against microsomal glutathione S-transferase.
Main Results:
- Plasma membranes exhibited significant membrane-bound glutathione S-transferase activity (83 +/- 8 mU/mg in canalicular, 50 +/- 3 mU/mg in sinusoidal).
- Plasma membrane GST activity showed a two-fold activation by N-ethylmaleimide, less than microsomal (seven-fold) and outer mitochondrial (four-fold) forms.
- Western blot confirmed a 17 kDa protein, recognized by the anti-microsomal GST antibody, in both membrane fractions, with higher reactivity in canalicular membranes.
Conclusions:
- Rat liver plasma membranes possess distinct glutathione S-transferase activity.
- The enzyme in plasma membranes is closely related to, but not identical to, microsomal and outer mitochondrial glutathione S-transferases.
- These findings suggest a specific role for plasma membrane-associated GST in liver cell function and detoxification.