Previous studies from this laboratory have established that acquired resistance of murine
L1210 leukemia cells to
L-phenylalanine mustard (
L-PAM) and other
alkylating agents is accompanied by a two-to threefold elevation in their
glutathione (GSH) concentration (Biochem. Pharm. 31:121). In an attempt to gain insight into the mechanism by which resistant
tumor cells maintain their increased GSH content, we have assessed the possible role of
gamma-glutamyl transpeptidase (gamma-GT), a membrane bound
enzyme involved in GSH metabolism. These results indicate that the
enzyme is present in both sensitive and resistant murine
L1210 leukemia cells but that the cellular content of gamma-GT is elevated two-to threefold in
L-PAM resistant cells as compared to their sensitive counterparts. This elevation in enzymatic activity correlates well with the increased cellular GSH content in resistant cells. The results of a detailed kinetic analysis of gamma-GT activity indicate that there is no difference, between cell types, in the apparent Km of the
enzyme for the gamma-glutamyl donor (
L-gamma-glutamyl-p-nitroanilide) or the acceptor (
glycylglycine). However, the apparent Vmax is increased two-to threefold in
L-PAM resistant
tumor cells. Investigation into the role of gamma-GT in the extracellular metabolism of GSH indicates that resistant
tumor cells metabolize two-fold more GSH than do sensitive cells and that such metabolism results in a similar difference in the intracellular concentration of
cysteine. Results of studies with cellular lysates also indicate a role for the
enzyme in the supply of
cysteine to the
glutathione precursor pool of the
tumor cell and in the maintenance of elevated GSH concentrations in cells resistant to
alkylating agents.