Characterization of the putative alpha subunit of a heterotrimeric G protein in rice

Plant Mol Biol. 1997 Jul;34(4):563-72. doi: 10.1023/a:1005807010811.

Abstract

A recombinant protein with a cDNA that encodes the putative alpha subunit of a rice heterotrimeric G protein was synthesized in Escherichia coli and purified. The recombinant protein (rGrice alpha) with an apparent molecular mass of 45 kDa was bound with guanosine 5'-(3-O-thio)triphosphate with an apparent association constant (kapp) of 0.36. The protein also hydrolyzed GTP and its kcat was 0.44. rGrice alpha was ADP-ribosylated by activated cholera toxin. Monoclonal antibodies raised against rGrice alpha reacted with a 45 kDa polypeptide localized in the plasma membrane of rice seedlings. The peptide map of this polypeptide after digestion with V8 protease was identical to that of rGrice alpha. A 45 kDa polypeptide in the plasma membrane, as well as rGrice alpha, was ADP-ribosylated by activated cholera toxin. The GTPase activity of the plasma membrane was stimulated 2.5-fold by mastoparan 7 but not mastoparan 17. These properties were similar to those of the alpha subunits of heterotrimeric G proteins in animals, suggesting that the putative alpha subunit is truly the alpha subunit itself.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine Diphosphate Ribose / metabolism
  • Antibody Specificity
  • Cell Compartmentation
  • Cell Fractionation
  • Cell Membrane / chemistry
  • Cell Membrane / immunology
  • Cholera Toxin / metabolism
  • DNA, Complementary / genetics
  • Escherichia coli / genetics
  • GTP-Binding Proteins / chemistry*
  • GTP-Binding Proteins / genetics
  • GTP-Binding Proteins / immunology
  • GTP-Binding Proteins / metabolism
  • GTPase-Activating Proteins*
  • Guanosine 5'-O-(3-Thiotriphosphate) / metabolism
  • Guanosine Triphosphate / metabolism
  • Oryza / chemistry
  • Oryza / genetics
  • Peptide Mapping
  • Plant Proteins / chemistry*
  • Plant Proteins / genetics
  • Plant Proteins / immunology
  • Plant Proteins / metabolism
  • Protein Conformation
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / immunology
  • Recombinant Proteins / metabolism

Substances

  • DNA, Complementary
  • GTPase-Activating Proteins
  • Plant Proteins
  • Recombinant Proteins
  • rho GTPase-activating protein
  • Adenosine Diphosphate Ribose
  • Guanosine 5'-O-(3-Thiotriphosphate)
  • Guanosine Triphosphate
  • Cholera Toxin
  • GTP-Binding Proteins