Posttranslationally processed structure of the human platelet protein smg p21B: evidence for geranylgeranylation and carboxyl methylation of the C-terminal cysteine

Proc Natl Acad Sci U S A. 1990 Nov;87(22):8960-4. doi: 10.1073/pnas.87.22.8960.

Abstract

smg p21A and -B are small GTP-binding proteins that share putative effector and consensus C-terminal sequences with ras p21 proteins. In the present report, we showed that human platelet smg p21B became labeled when intact platelets were incubated with exogenous [3H]mevalonolactone and when a purified preparation of smg p21B was incubated with bovine brain membranes and S-adenosyl-L-[methyl-3H]methionine. In addition, we demonstrated by gas chromatography/mass spectrometry that treatment of smg p21B with Raney nickel released a geranylgeranyl moiety in a molar ratio of about 1:1. In contrast, treatment of smg p21B with NH2OH or KOH yielded no evidence for the presence of a palmitoyl thioester. Extensive digestion of smg p21B with Achromobacter protease I yielded two C-terminal tripeptides that contained serine and cysteine in a molar ratio of 2:1. Both peptides were modified by a thioether-linked geranylgeranyl group. One of the peptides comigrated with a 3H-labeled proteolytic product of methylated smg p21B on reverse-phase HPLC and this peptide appeared at the same retention time as that of the other peptide after being treated with KOH. Since the cDNA-predicted C-terminal sequence of smg p21B contains a unique Ser-Ser-Cys peptide within its C-terminal domain, -Lys-Lys-Ser-Ser-Cys-Gln-Leu-Leu184, these results indicate that smg p21B is posttranslationally modified by geranylgeranylation of Cys-181 and suggest that further modifications cause proteolytic removal of the three predicted C-terminal amino acids followed by partial methylation of the cysteinyl carboxyl group.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Blood Platelets / metabolism*
  • Cysteine / metabolism
  • GTP-Binding Proteins / chemistry
  • GTP-Binding Proteins / metabolism*
  • Gas Chromatography-Mass Spectrometry
  • Humans
  • In Vitro Techniques
  • Methylation
  • Mevalonic Acid / analogs & derivatives
  • Mevalonic Acid / chemistry
  • Polyisoprenyl Phosphates / metabolism*
  • Protein Processing, Post-Translational*
  • rap GTP-Binding Proteins

Substances

  • Polyisoprenyl Phosphates
  • mevalonolactone
  • GTP-Binding Proteins
  • rap GTP-Binding Proteins
  • Cysteine
  • Mevalonic Acid