Identification of gamma-glutamyltransferase in rat liver plasma membranes after two-dimensional electrophoresis
Two-dimensional electrophoresis and sodium dodecyl sulfate-polyacrylamide gel electrophoresis, followed by silver staining or specific immunoblotting analysis, was applied to the characterization of gamma-glutamyltransferase (GGT, EC 2.3.2.2.)in rat tissues. We confirmed that purified kidney GGT is a dimer composed of two nonidentical subunits with molecular masses of about 50 and 30 kDa. Both thelight and the heavy subunits were separated into 6 and 12 protein bands, respectively. Antibody fractions against the 30 and 50 kDa subunits, purified by immunoaffnity from a whole antiserum directed to the rat kidney enzyme, recognized their corresponding subunits and did not cross-react with each other. Studies of the reactivity of these antibodies towards GGTs from kidney and liver homogenates revealed an intraspecies dissimilarity in the molecular architecture of the kidney and liver GGTs, especially concerning the 30 kDa subunit. Following phenobarbital treatment of animals we observed an increase in immunoreactive GGT, accompanied by the appearance of additional polypeptides with more basic isoelectric points and higher molecular mass.