Geranylgeranylated Rab proteins terminating in Cys-Ala-Cys, but not Cys- Cys, are carboxyl-methylated by bovine brain membranes in vitro

Tor E. Smeland, Miguel C. Seabra, Joseph L. Goldstein, Michael S. Brown

Research output: Contribution to journalArticle

62 Scopus citations

Abstract

Geranylgeranylated Rab proteins usually terminate in either Cys-Cys or Cys-Xaa-Cys, where Xaa is Ala, Ser, or Gly. In both classes of proteins, the two cysteines are geranylgeranylated, but only the Cys-Xaa-Cys class has been shown to be carboxyl-methylated on the terminal cysteine in vivo. In the current study, we used recombinant Rab geranylgeranyltransferase and a Rab escort protein (REP-1) to attach geranylgeranyl residues to the two cysteines at the carboxyl terminus of Rab3A (Cys-Ala-Cys) and Rab1A (Cys-Cys). The geranylgeranylated proteins were then incubated with bovine cerebellar membranes that contain an enzyme that transfers [3H]methyl from S-[methyl- 3H]adenosyl-L-methionine to geranylgeranylated cysteine. The enzyme transferred [3H]methyl to geranylgeranylated Rab3A but not to geranylgeranylated Rab1A. Replacement of the Cys-Ala-Cys terminus of Rab3A with Cys-Cys abolished methylation, and the opposite result was obtained when the Cys-Cys of Rab1A was replaced with Cys-Ala-Cys. When the Cys-Cys terminus of Rab1A was changed to Ser-Cys, methylation was restored. These studies suggest that the carboxyl-terminal cysteine of Rab proteins terminating in Cys-Xaa-Cys but not Cys-Cys is methylated and that the resistance of Cys-Cys proteins to methylation is attributable to the vicinal geranylgeranylated cysteines.

Original languageEnglish (US)
Pages (from-to)10712-10716
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume91
Issue number22
DOIs
StatePublished - Nov 7 1994

Keywords

  • Rab geranylgeranyltransferase
  • carboxyl methyltransferase
  • membrane traffic
  • protein prenylation

ASJC Scopus subject areas

  • General

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