SUBSTITUTION OF CADMIUM FOR ZINC IN FARNESYL-PROTEIN TRANSFERASE ALTERS ITS SUBSTRATE-SPECIFICITY

Ras proteins are mutationally activated in a variety of human cancers. Since farnesylation of Ras proteins is required for expression of the ir oncogenic potential. the enzyme responsible for this reaction, farn esyl:protein transferase (FPT), has become a major target for anticanc er drug development. FPT is a zinc metalloenzyme, and the zinc is esse ntial for its catalytic activity. To begin to elucidate the role of zi nc in catalysis, we initiated metal substitution studies. Of all metal s tested, only cadmium was able to functionally substitute for zinc, r econstituting enzymatic activity with native substrates (H-Ras and far nesyl diphosphate) to about 50% of that of the zinc-containing enzyme. Several important differences were observed between cadmium-substitut ed FPT (Cd-FPT) and zinc-containing FPT (Zn-FPT). Cd-FPT not only uses H-ras with its native CaaX motif (Ras-CVLS) as a substrate but also f an farnesylate H-ras in which the CaaX motif is altered to contain a C -terminal leucine residue (Ras-CVLL). Similarly, Cd-FPT can farnesylat e leucine-terminated peptides. Leucine-terminated proteins and peptide s are usually substrates for the related enzyme geranylgeranyl:protein transferase type I. Farnesylation of Ras-CVLS and Res CVLL by Cd-FPT exhibited similar sensitivity to the FPT inhibitor SCH 44342 and to th e peptide inhibitor CAIM. However, unlike Zn-FPT, Cd-FPT is also poten tly inhibited by the leucine-terminated peptide CAIL. These results in dicate that the metal ion content of FPT strongly influences its prote in substrate specificity.