|Budget Amount *help
¥2,400,000 (Direct Cost: ¥2,400,000)
Fiscal Year 1997: ¥900,000 (Direct Cost: ¥900,000)
Fiscal Year 1996: ¥1,500,000 (Direct Cost: ¥1,500,000)
One of the unanswered questions in this project is the presumed role that prenylation plays in targeting many proteins to specific cellular membranes. While ras and most heterotrimeric G proteins are specifically localized to the plasma membrane, other prenylated proteins, such as members of the rab family, are predominantly localized on intracellular membranes. Scince these differences in localization cannot be accounted for by distinct isoprenoids (farnesylation or geranylgeranylation), additional determinants must be involved. One possibility is that specific receptors on membranes would recognize both the isoprenoid and an additional region of a particular protein. Another possibility is that a prenylated protein could be recognized by a putative molecular chaperon which would direct it to a specific membrane. Elucidating the molecular mechanisms should provide new insight into the importance of prenylation in intracellular signal transduction by isoprenylated proteins. We report the identification, purification, and characterization of proteins that interact with farnesyl moiety of ras protein. The proteins were purified from rat brain cytosol through use of an affinity column containing the farnesylated heptapeptide (SKTKC) corresponding to the COOH-terminal 5 amino acids of Ki-ras. Major two proteins were purified by this chromatography. One has an apparent molecular weight of 50,000, termed P50, and that of another protein (P90) is 90,000 on SDS-polyacrylamide gel electrophoresis. Sequence analysis of lysyl endopeptidase-released peptides from these proteins revealed that P50 is a mixture of alpha, beta-tubulin and P90 is a mixture of alpha, beta-heat shock proteins 90 (hsp90). Further study is required to elucidate physiological function of these proteins interacting with the farnesyl moiety of ras protein.