MOLECULAR-CLONING, CHARACTERIZATION, AND EXPRESSION IN ESCHERICHIA-COLI OF FULL-LENGTH CDNA OF 3 HUMAN GLUTATHIONE-S-TRANSFERASE-PI GENE VARIANTS - EVIDENCE FOR DIFFERENTIAL CATALYTIC ACTIVITY OF THE ENCODED PROTEINS

We report the isolation of three full-length cDNAs corresponding to th e mRNAs of closely related glutathione S-transferase (GST) Pi genes, d esignated hGSTP1A, hGSTP1*B, and hGSTP1*C, expressed in normal cells and malignant gliomas. The variant cDNAs result from A --> G and C --> T transitions at nucleotides +313 and +341, respectively. The transit ions changed codon 104 from ATC (Ile) in hGSTP1A to GTC (Val) in hGST P1B and h-GSTP1*C and changed codon 113 from GCG (Ala) to GTG (Val) i n hGSTP1C. Both amino changes are in the electrophile-binding active site of the GST Pi peptide. Computer modeling of the deduced crystal s tructures of the encoded peptides showed significant deviations in the interatomic distances of critical electrophile-binding active site am ino acids as a consequence of the amino acid changes. The encoded prot eins expressed in Escherichia coli and purified by GSH affinity chroma tography showed a 3-fold lower K-m (CDNB) and a 3-4-fold higher K-cat/ K-m for the hGSTP1A encoded protein than the proteins encoded by hGST P1B and hGSTP1*C. Analysis of 75 cases showed the relative frequency of hGSTP1C to be 4-fold higher in malignant gliomas than in normal ti ssues. These data provide conclusive molecular evidence of allelopolym orphism of the human GST Pi gene locus, resulting in active, functiona lly different GST Pi proteins, and should facilitate studies of the ro le of this gene in xenobiotic metabolism, cancer, and other human dise ases.