ENZYMATIC CHARACTERISTICS OF CHIMERIC MYC RYC(1) GLUTATHIONE S-TRANSFERASES/

Mice are resistant to aflatoxin carcinogenicity primarily due to expre ssion of a glutathione S-transferase (mYc) with high catalytic activit y toward aflatoxin B-1-8,9-epoxide (AFBO). In contrast, rats are more sensitive to aflatoxin carcinogenicity due to the constitutive express ion of a glutathione S-transferase with relatively low catalytic activ ity toward AFBO (rYc(1)). To identify the contribution of different re gions of the mYc protein that confer high catalytic activity toward AF BO, six chimeric mYc/rYc(1) GST enzymes were generated utilizing full and partial restriction enzyme digestions at two conserved StyI sites- in the mYc and rYc(1) complementary DNAs (between amino acid residues 56-57 and 142-143). Recombinant wild-type and chimeric glutathione S-t ransferases mere bacterially expressed, affinity purified, and their c atalytic activities measured toward AFBO, Delta(5)-androstene-3,17-dio ne, 1-chloro-2,4-dinitrobenzene, and ethacrynic acid. The set of chime ras displayed a wide range of catalytic activities toward the substrat es assayed. The chimeras with the greatest activity toward AFBO were 1 :56rat-57: 221mouse and 1:56mouse-57:142rat-143:221mouse, with APBO co njugating activities 200 and 8 times greater than wild-type rYc(1),res pectively. These results demonstrate that the residues that confer hig h AFBO conjugation activity in mYc are located in the region spanning residues 57-221.