Preliminary Characterization of Glutathione S-Transferases That Accumulate in Callus Cells of Pumpkin (Cucurbita maxima Duch.)

Glutathione S-transferases (GSTs; EC 2.5.1.18 EC] ) in sarcocarptissue of pumpkin (Cucurbita maxima Duch.) fruit and in callusinduced from the tissue were examined. The specific activityof GST in the callus was 6.9-fold higher than that in the tissue.The specific activity in the callus remained constant duringcultivation. Column chromatography on DEAE-cellulose, hydroxylapatite,and S-hexylglutathione-agarose was used to fractionate solubleproteins that were precipitated by ammonium sulfate at 30% to70% saturation from homogenates of the sarcocarp tissue of pumpkinfruit and the callus and GST activity was monitored. Two andseven isozymes of GST were identified in the tissue and in thecallus, respectively. Furthermore, column chromatography onSephadex G-200 and SDS-polyacrylamide gel electrophoresis, indicatedthat these GST isozymes were homo- and heterodimers of subunitsof M_r 22,000 (Puga), and 23,000 (Pugb), 24,000 (Pugc) or 24,500(Pugd). Puga and Pugb were predominant in the sarcocarp tissueand in the callus, respectively. Puga, Pugb, Pugc and Pugd hadacidic pI values of 5.45, 5.00, 5.35 and 5.75, respectively.Rabbit antiserum against Pugb did not cross-react with the threeother subunits of GST during immunoblotting. (Received July 15, 1993; Accepted December 14, 1993)