Abstract
Native glutathione S-transferases are composed of subunits with apparent molecular weights of 25,000, 23,500, or 22,000 which form either homo- or heterodimers. Glutathione S-transferases A, C, and X which contain two subunits with molecular weights of 23,500 yielded similar but nonidentical proteolytic fragmentation patterns. Fragments unique to the subunits of the homodimers A and X were present in decreased intensities in the patterns of form C. Two-dimensional electrophoresis under denaturing conditions showed single nonoverlapping spots for transferases A and X, while form C yielded two spots corresponding in position to those obtained from forms A and X. Renaturation of dissociated glutathione S-transferase C yielded enzymatically active transferases A, C, and X. These results indicate that form C is a heterodimer composed of one subunit from the homodimeric transferases A and X. This was substantiated by NH2-terminal sequence analysis showing extensive NH2-terminal homology amongst all three forms. However, in the positions where forms A and X yielded different residues, both amino acids were detected in the sequence of form C, indicating that the two subunits of Mr = 23,500 are the products of two different genes. NH2-terminal sequence analysis of the heterodimeric glutathione S-transferase B which is composed of subunits with molecular weights of 22,000 and 25,000 revealed a single unique sequence which bore no resemblance to the sequences of either forms A or X. Despite the identical NH2-terminal sequences, proteolytic fragmentation of the separated subunits showed markedly different fragmentation patterns. This indicates that two different mRNAs code for these two subunits.
Highlights
23,500, or 22,000 which form either homo- or heter- (Litwack et al, 1971; Hayes et al, 1979), and X which has odimers
Native glutathione S-transferases acroemposed of subunits with apparentmolecular weights of 25,000,23,500, and 22,000 which exist as either homo- or heterodimers (Hayes et al, 1980).Glutathione S-transferaseB is a heterodimecromposed of the subunits with molecular weights of 25,000 and 22,000, while glutathione S-transferase AA and ligandin are homodimers with subunit molecular weights of 25,000 and 22,000, respectively (Kithara and Sato,1981)
The two subunits forming the glutathione S-transferases A, C, and X are indistinguishable in molecular weight (Mr= 23,500) and share antigenic determinants, they mology amongst all three forms
Summary
Undenatured glutathione S-transferase C as well as and X-The subunit composition of the transferases A, C, glutathione S-transferases A, C, and X treated with urea and renaand X was investigated by two-dimensional gel electrophoresis under denaturing conditions (Fig. 1). In this system the transferases A and X each yielded one major spot, with turcd in the absenocfe urea were subsequently subjected to isoelectric focusing under nondenaturing conditions, and the enzyme activity was determined in gel fractions. The isoelectric and X-Experiments described above (Fig. 2 ) suggested that time "- Subunit Composition of GlutathioneS-Transferases (min) chyme + + + + - + + + + - + + + + -
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