Isoelectric focusing of glutathione S-transferases from rat liver and kidney

Abstract

The glutathione S-transferases [EC 2.5.1.18] that were purified to homogeneity from liver cytosol have overlapping but distinct substrate specificities and different isoelectric points [PI]. This report explores the possibility of using preparative electrofocusing to compare the composition of the transferases in liver and kidney cytosol. Hepatic cytosol from adult male Sprague-Dawley rats was resolved by isoelectric focusing on Sephadex columns into 5 peaks of transferase activity, each with characteristic substrate specificity. The first 4 peaks of transferase activity (in order of decreasing basicity) are identified as transferases AA, B, A and C, respectively, on the basis of substrate specificity, but the 5th peak (pI 6.6) does not correspond to a previously described transferase. Isoelectric focusing of renal cytosol resolves only 3 major peaks of transferase activity, each with narrow substrate specificity. In the kidney, peak 1 (pI 9.0) has most of the activity toward 1-chloro-2,4-dinitrobenzene, peak 2 (pI 8.5) toward p-nitrobenzyl chloride and peak 3 (pI 7.0) toward trans-4-phenylbut-3-en-2-one. Renal transferase peak 1 (pI 9.0) appears to correspond to transferase B on the basis of pI, substrate specificity and antigenicity. Kidney transferase peaks 2 (pI 8.5) and 3 (pI 7.0) do not correspond to previously described glutathione S-transferases, although kidney transferase peak 3 is similar to the transferase peak 5 from focused hepatic cytosol. Transferases A and C were not found in kidney cytosol, and transferase AA was detected in only 1 out of 6 replicates. Thus it is important to recognize the contribution of individual transferases to total transferase activity in that each transferase may be regulated independently.