Characterization of a guanine nucleotide-releasing factor and a GTPase-activating protein that are specific for the ras-related protein p25rab3A

Proc Natl Acad Sci U S A. 1992 Feb 15;89(4):1154-8. doi: 10.1073/pnas.89.4.1154.

Abstract

The rab3A gene product is a 25-kilodalton guanine nucleotide-binding protein that is expressed at high levels in neural tissue and has about 30% homology to the ras gene product. Recombinant Rab3A protein and p25rab3A purified from bovine brain membranes have been used as substrates to look for factors that regulate its biochemical activity. A factor in rat brain cytosol exists that accelerates, by approximately 10-fold, the release and subsequent rebinding of guanine nucleotides to both native and recombinant p25rab3A. We have partially purified this activity, termed Rab3A-GRF, and a GTPase-activating protein (Rab3A-GAP) reported previously. The two activities copurified through a variety of procedures but were separated by Mono Q anion-exchange chromatography, indicating that the activities arise from distinct polypeptides. Both factors were thermolabile, sensitive to trypsin, and specific for Rab3A, exhibiting little or no activity toward c-Ha-Ras or Rab2 proteins. By gel filtration chromatography and sucrose density ultracentrifugation, both Rab3A-GRF and Rab3A-GAP have Stokes radii of 79 A and sedimentation coefficients of 8.9 S. We calculate a molecular mass of 295,000 daltons and a frictional ratio of 1.80 for each factor.

Publication types

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

MeSH terms

  • Animals
  • Brain Chemistry
  • GTP-Binding Proteins / isolation & purification
  • GTP-Binding Proteins / physiology*
  • GTPase-Activating Proteins
  • Guanine Nucleotides / metabolism*
  • Nerve Tissue Proteins / physiology*
  • Proteins / physiology*
  • Rats
  • rab3 GTP-Binding Proteins
  • ras GTPase-Activating Proteins

Substances

  • GTPase-Activating Proteins
  • Guanine Nucleotides
  • Nerve Tissue Proteins
  • Proteins
  • ras GTPase-Activating Proteins
  • GTP-Binding Proteins
  • rab3 GTP-Binding Proteins