GTP-stimulated carboxyl methylation of a soluble form of the GTP-binding protein G25K in brain.
The GTP-stimulated carboxyl methylation of an M(r) 23,000 protein was investigated in brain homogenates. An M(r) 23,000 methylation substrate was purified from brain homogenates, using an assay for protein methyl-acceptor activity in the presence of a membrane-bound methyltransferase. The M(r) 23,000 methyl-acceptor protein was identified as a soluble form of the GTP-binding protein G25K, based on antibody reactivity and amino acid sequences of tryptic peptides. Two forms of methylated G25K, differing in isoelectric points, were isolated. The soluble G25K could be methylated with a stoichiometry approaching 1 mol of methyl group per mol of G25K, and guanosine 5'-O-3-(thio)triphosphate stimulated the methylation by decreasing the Km for G25K from 0.79 to 0.17 microM. After methylation, the G25K was associated with the membrane fraction. The soluble G25K was isolated as a heterodimer of G25K and an M(r) 28,000 protein. The G25K and M(r) 28,000 protein complex was dissociated with 3-[(3-cholamidopropyl)-dimethylammonio]-1-propanesulfonate detergent, and the subunits were separated by Mono-Q chromatography. The association of the M(r) 28,000 protein with G25K decreased the methylation of G25K and altered the guanine nucleotide specificity, indicating that the M(r) 28,000 protein may regulate the methylation of G25K.
Backlund
P S
PS
Laboratory of General and Comparative Biochemistry, National Institute of Mental Health, Bethesda, Maryland 20892.
D000818:Animals; D015153:Blotting, Western; D002850:Chromatography, Gel; D015180:Electrophoresis, Gel, Two-Dimensional; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006150:Guanine Nucleotides / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism*; D008745:Methylation; D011257:Pregnancy Proteins / Q000378:metabolism*; D011817:Rabbits; D020764:cdc42 GTP-Binding Protein
D006150:Guanine Nucleotides; D011257:Pregnancy Proteins; D006160:Guanosine Triphosphate; D019204:GTP-Binding Proteins; D020764:cdc42 GTP-Binding Protein