Purification and characterization of a GTP-binding protein with a molecular weight of 20,000 in bovine brain membranes. Identification as the rho gene product.

摘要: Abstract We have determined that there are at least six GTP-binding proteins (G proteins) with Mr values between 20,000 and 25,000 in the crude membrane fraction of bovine brain purified one them a about 24,000 (24K G) to near homogeneity (Kikuchi, A., Yamashita, T., Kawata, M., Yamamoto, K., Ikeda, Tanimoto, Takai, Y. (1988) J. Biol. Chem. 263, 2897-2904). In this study, we another G protein (20K characterized it. 20K bound maximally 1.0 mol [35S]guanosine 5'-(3-O-thio)triphosphate (GTP gamma S)/mol protein, Kd value 50 nM. [35S]GTP S binding was inhibited by GTP GDP, but not other nucleotides such as ATP, UTP, CTP; it also pretreatment N-ethylmaleimide. hydrolyzed liberate Pi, turnover number 0.01 min-1, copurified beta subunits regulatory adenylate cyclase. recognized antibody against ADP-ribosylation factor for stimulatory Peptide map analysis showed proteolytic product 24K G. The partial amino acid sequence almost identical deduced from rho gene. composition similar gene product. These results suggest is present membranes.