Differences in stereoselectivity and catalytic efficiency of three human glutathione transferases in the conjugation of glutathione with 7 beta,8 alpha-dihydroxy-9 alpha,10 alpha-oxy-7,8,9,10-tetrahydrobenzo(a)pyrene.

The kinetics of the enzyme-catalyzed conjugation of glutathione with (+/-)-7 beta, 8 alpha-dihydroxy-9 alpha, 10 alpha -oxy-7,8,9,10 -tetrahydrobenzo(a)pyrene [(+/-)-anti-BPDE] have been studied with the following human cytosolic glutathione transferases: the basic (alpha-epsilon) and near-neutral (mu) isoenzymes from liver, and the acidic (pi) isoenzyme from placenta. When the BPDE concentration was varied (using 5 mM glutathione) the apparent Vmax values for transferases alpha-epsilon, mu, and pi were 38, 570, and 825 nmol X mg-1 X min-1, respectively, with corresponding apparent Km values of 88, 27, and 54 microM. The apparent Km values for glutathione [using 80 microM (+/-)-anti-BPDE] were 0.4, 0.7, and 0.1 mM for transferase alpha-epsilon, mu, and pi, respectively. The glutathione conjugates formed with the two enantiomers of (+/-)-anti-BPDE were resolved by high performance liquid chromatography. The percentages of conjugates derived from the highly carcinogenic (+)-enantiomer were 59, 60, and greater than or equal to 90% for transferases alpha-epsilon, mu, and pi, respectively. The separate enantiomers of anti-BPDE were assayed in experiments with transferases mu and pi. Both enantiomers were substrates for transferase mu, but only the (+)-enantiomer gave measurable activity with transferase pi. A 3-fold increase in Vmax and Km values for transferase pi was obtained with (+)-anti-BPDE as compared with the racemic substrate and could be quantitatively accounted for by the finding that (-)-anti-BPDE serves as a competitive inhibitor for transferase pi.