AMINO-ACID DIFFERENCES AT POSITION-10, POSITION-11, AND POSITION-104 EXPLAIN THE PROFOUND CATALYTIC DIFFERENCES BETWEEN 2 MURINE PI-CLASS GLUTATHIONE S-TRANSFERASES

The glutathione S-transferases play a pivotal role in the detoxificati on of toxic and carcinogenic electrophiles. We have previously reporte d the isolation of two actively transcribed murine pi-class glutathion e S-transferase genes. In this study the two proteins encoded by these genes, Gst p-1 and Gst p-2, were expressed in Escherichia coli and fo und to exhibit profoundly different catalytic activities, the activity of Gst p-2 toward a panel of electrophilic substrates being 1-3 order s of magnitude lower than that of Gst p-1. In. order to establish the basis for the difference between these highly homologous proteins, mut ants were generated where specific amino acids had been exchanged. Kin etic analysis of the wild-type and mutant enzymes revealed that the am ino acid differences occurring at positions 10 (Val/Ser), 11 (Arg/Pro) , and 104 (Val/Gly) are responsible for the reduced enzymatic activity of Gst p-2. This analysis together with computer graphics modeling fo r Gst p-2 indicated that these changes affected both substrate and glu tathione binding to the enzyme.