In the yeast Saccharomyccs cerevwise, qiutations in either of two unlinked genes, RAM)-1or RAM2, abolish the farnesyltrnsferae activity responsible for prenylation of Ras proteins and the a-factor mating pheromone. Here we report that thefunction of RAM) antd RAM2 genme is required for the membrane laliation ofRas proteins and a-factor. The RAM2 gene was sequenced and can encode a 38-kDa protein.We examined tie functional interaction ofRAM2 andRAMf by expressing the genes in Escherchia colt. Extracts derived from an E. coil strain that coexpressed RAM] and RAM2 efficiently farnesylated a-factor peptide and Ras protein substrates. In contrast, extracts derived from E. colN strains that expressed either RAM gene alone were devoid of activity; however, when the latter extracts were mixed, protein farnesyltransferaSe activity was reconstituted. These results indicate that the yeast farnesyl-protein transferase is comprised of Raml and Ram2 polypeptides. Although Raml is a component of the enzyme, disruption of the RAM) gene in yeast was not lethal indicating that the Raml-Ram2 farnesyltserase is not essential for viability. In contrast, disruption of RAM2 was lethal, suggesting that Ram2 has an essential function in addition to its role with Raml in protein farnesylation.The post-translational modification of proteins by the covalent attachment of isoprenoid groups plays an important role in the membrane targeting of various proteins (1-5). Three classes of prenylated proteins in eukaryotic cells have been described. The first class, represented by certain fungal mating pheromones, Ras proteins, and nuclear lamins, is initially synthesized with a C-terminal CAAX sequence, where X = Ser, Cys, Met, or Ala, and is modified by thioether linkage of a C15 farnesyl group to the cysteine residue (6-9). The second class of prenylated proteins, represented by the y subunits of certain heterotrimeric G proteins and various Ras-like proteins, is initially synthesized with different C-terminal CAAX sequences, in which X = Leu or Phe, and is modified by the attachment of a C20 geranylgeranyl group to the cysteine residue (10-12). Members of both of these classes of prenylated proteins are subsequently processed by proteolytic removal of the three terminal amino acids and methylation of the newly exposed carboxyl group of the prenyl-cysteine (6, 10-15). The third class of prenylated proteins, represented by the Yptl/Sec4 (Rab) family of Ras-like GTPases that terminate with the sequence CC or CXC, is also geranylgeranylated at a C-terminal cysteine residue(s) (16)(17)(18).

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