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Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1995 May 23;92(11):5008–5011. doi: 10.1073/pnas.92.11.5008

A mechanism for posttranslational modifications of proteins by yeast protein farnesyltransferase.

J M Dolence 1, C D Poulter 1
PMCID: PMC41837  PMID: 7761439

Abstract

Protein farnesyltransferase catalyzes the alkylation of cysteine in C-terminal CaaX sequences of a variety of proteins, including Ras, nuclear lamins, large G proteins, and phosphodiesterases, by farnesyl diphosphate (FPP). These modifications enhance the ability of the proteins to associate with membranes and are essential for their respective functions. The enzyme-catalyzed reaction was studied by using a series of substrate analogs for FPP to distinguish between electrophilic and nucleophilic mechanisms for prenyl transfer. FPP analogs containing hydrogen, fluoromethyl, and trifluoromethyl substituents in place of the methyl at carbon 3 were evaluated as alternative substrates for alkylation of the sulfhydryl moiety in the peptide dansyl-GCVIA. The analogs were alternative substrates for the prenylation reaction and were competitive inhibitors against FPP. A comparison of kcat for FPP and the analogs with ksolv, the rate constants for solvolysis of related p-methoxybenzenesulfonate derivatives, indicated that protein prenylation occurred by an electrophilic mechanism.

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Selected References

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  1. Armstrong S. A., Seabra M. C., Südhof T. C., Goldstein J. L., Brown M. S. cDNA cloning and expression of the alpha and beta subunits of rat Rab geranylgeranyl transferase. J Biol Chem. 1993 Jun 5;268(16):12221–12229. [PubMed] [Google Scholar]
  2. Ashby M. N., Edwards P. A. Elucidation of the deficiency in two yeast coenzyme Q mutants. Characterization of the structural gene encoding hexaprenyl pyrophosphate synthetase. J Biol Chem. 1990 Aug 5;265(22):13157–13164. [PubMed] [Google Scholar]
  3. Barbacid M. ras genes. Annu Rev Biochem. 1987;56:779–827. doi: 10.1146/annurev.bi.56.070187.004023. [DOI] [PubMed] [Google Scholar]
  4. Bos J. L. ras oncogenes in human cancer: a review. Cancer Res. 1989 Sep 1;49(17):4682–4689. [PubMed] [Google Scholar]
  5. Brown M. S., Goldstein J. L., Paris K. J., Burnier J. P., Marsters J. C., Jr Tetrapeptide inhibitors of protein farnesyltransferase: amino-terminal substitution in phenylalanine-containing tetrapeptides restores farnesylation. Proc Natl Acad Sci U S A. 1992 Sep 1;89(17):8313–8316. doi: 10.1073/pnas.89.17.8313. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Cassidy P. B., Dolence J. M., Poulter C. D. Continuous fluorescence assay for protein prenyltransferases. Methods Enzymol. 1995;250:30–43. doi: 10.1016/0076-6879(95)50060-x. [DOI] [PubMed] [Google Scholar]
  7. Clarke S. Protein isoprenylation and methylation at carboxyl-terminal cysteine residues. Annu Rev Biochem. 1992;61:355–386. doi: 10.1146/annurev.bi.61.070192.002035. [DOI] [PubMed] [Google Scholar]
  8. Der C. J., Cox A. D. Isoprenoid modification and plasma membrane association: critical factors for ras oncogenicity. Cancer Cells. 1991 Sep;3(9):331–340. [PubMed] [Google Scholar]
  9. Farnsworth C. C., Kawata M., Yoshida Y., Takai Y., Gelb M. H., Glomset J. A. C terminus of the small GTP-binding protein smg p25A contains two geranylgeranylated cysteine residues and a methyl ester. Proc Natl Acad Sci U S A. 1991 Jul 15;88(14):6196–6200. doi: 10.1073/pnas.88.14.6196. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Gomez R., Goodman L. E., Tripathy S. K., O'Rourke E., Manne V., Tamanoi F. Purified yeast protein farnesyltransferase is structurally and functionally similar to its mammalian counterpart. Biochem J. 1993 Jan 1;289(Pt 1):25–31. doi: 10.1042/bj2890025. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Marshall C. J. Protein prenylation: a mediator of protein-protein interactions. Science. 1993 Mar 26;259(5103):1865–1866. doi: 10.1126/science.8456312. [DOI] [PubMed] [Google Scholar]
  12. Mayer M. P., Prestwich G. D., Dolence J. M., Bond P. D., Wu H. Y., Poulter C. D. Protein farnesyltransferase: production in Escherichia coli and immunoaffinity purification of the heterodimer from Saccharomyces cerevisiae. Gene. 1993 Sep 30;132(1):41–47. doi: 10.1016/0378-1119(93)90512-2. [DOI] [PubMed] [Google Scholar]
  13. Moores S. L., Schaber M. D., Mosser S. D., Rands E., O'Hara M. B., Garsky V. M., Marshall M. S., Pompliano D. L., Gibbs J. B. Sequence dependence of protein isoprenylation. J Biol Chem. 1991 Aug 5;266(22):14603–14610. [PubMed] [Google Scholar]
  14. Omer C. A., Kral A. M., Diehl R. E., Prendergast G. C., Powers S., Allen C. M., Gibbs J. B., Kohl N. E. Characterization of recombinant human farnesyl-protein transferase: cloning, expression, farnesyl diphosphate binding, and functional homology with yeast prenyl-protein transferases. Biochemistry. 1993 May 18;32(19):5167–5176. doi: 10.1021/bi00070a028. [DOI] [PubMed] [Google Scholar]
  15. Pompliano D. L., Rands E., Schaber M. D., Mosser S. D., Anthony N. J., Gibbs J. B. Steady-state kinetic mechanism of Ras farnesyl:protein transferase. Biochemistry. 1992 Apr 21;31(15):3800–3807. doi: 10.1021/bi00130a010. [DOI] [PubMed] [Google Scholar]
  16. Pompliano D. L., Schaber M. D., Mosser S. D., Omer C. A., Shafer J. A., Gibbs J. B. Isoprenoid diphosphate utilization by recombinant human farnesyl:protein transferase: interactive binding between substrates and a preferred kinetic pathway. Biochemistry. 1993 Aug 17;32(32):8341–8347. doi: 10.1021/bi00083a038. [DOI] [PubMed] [Google Scholar]
  17. Poulter C. D., Satterwhite D. M. Mechanism of the prenyl-transfer reaction. Studies with (E)- and (Z)-3-trifluoromethyl-2-buten-1-yl pyrophosphate. Biochemistry. 1977 Dec 13;16(25):5470–5478. doi: 10.1021/bi00644a012. [DOI] [PubMed] [Google Scholar]
  18. Reiss Y., Goldstein J. L., Seabra M. C., Casey P. J., Brown M. S. Inhibition of purified p21ras farnesyl:protein transferase by Cys-AAX tetrapeptides. Cell. 1990 Jul 13;62(1):81–88. doi: 10.1016/0092-8674(90)90242-7. [DOI] [PubMed] [Google Scholar]
  19. Reiss Y., Stradley S. J., Gierasch L. M., Brown M. S., Goldstein J. L. Sequence requirement for peptide recognition by rat brain p21ras protein farnesyltransferase. Proc Natl Acad Sci U S A. 1991 Feb 1;88(3):732–736. doi: 10.1073/pnas.88.3.732. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Schaber M. D., O'Hara M. B., Garsky V. M., Mosser S. C., Bergstrom J. D., Moores S. L., Marshall M. S., Friedman P. A., Dixon R. A., Gibbs J. B. Polyisoprenylation of Ras in vitro by a farnesyl-protein transferase. J Biol Chem. 1990 Sep 5;265(25):14701–14704. [PubMed] [Google Scholar]
  21. Schafer W. R., Kim R., Sterne R., Thorner J., Kim S. H., Rine J. Genetic and pharmacological suppression of oncogenic mutations in ras genes of yeast and humans. Science. 1989 Jul 28;245(4916):379–385. doi: 10.1126/science.2569235. [DOI] [PubMed] [Google Scholar]
  22. Spector T., Cleland W. W. Meanings of Ki for conventional and alternate-substrate inhibitors. Biochem Pharmacol. 1981 Jan 1;30(1):1–7. doi: 10.1016/0006-2952(81)90277-x. [DOI] [PubMed] [Google Scholar]

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