Insulin stimulates GDP release from G proteins in the rat and human liver plasma membranes.
Plasma membranes (1-2 mg protein) prepared from the livers of adult male rats and human organ donors were incubated with 0.6 microM [alpha-32P] guanosine triphosphate (GTP) in an adenosine triphosphate (ATP)-regenerating buffer at 37 degrees C for 1 h; during this incubation, the [32P]GTP is hydrolyzed and the nucleotide that is predominantly bound to the membranes is [32P] guanosine diphosphate (GDP). [32P]GDP release from the liver membranes was proportional to the protein concentration and increased as a function of time. At 5 mM, Ca2+, Mg2+, Mn2+, and Zn2+ maximally inhibited GDP release by 80-90%, whereas, 5 mM Cu2+ maximally stimulated the reaction by 100%. Therefore, cations were not included in the buffer used in the GDP release step. One microM Gpp(NH)p (5'-guanylylimidodiphosphate), a nonhydrolyzable analog of GTP, maximally stimulated [32P]GDP release in the liver membranes by up to 30%. Although 10 nM Gpp(NH)p had no effect on GDP release, it appeared to stabilize the hormonal effect by blocking further GDP/GTP exchange. In the rat membranes, 1-100 nM glucagon (used as a positive control) stimulated [32P]GDP release by about 17% (P < .05); similarly, 0.1-100 nM insulin stimulated [32P]GDP release by 10-13% (P < .05). In the human membranes, 10 pM to 100 nM insulin stimulated [32P]GDP release by 7-10%. In the rat membranes, 10 nM insulin stimulated [32P]GDP release by 17 and 24% at 2 and 4 min, respectively (P < .05); in the human membranes, 10 nM insulin stimulated [32P]GDP release by about 9% at 2 and 4 min.(ABSTRACT TRUNCATED AT 250 WORDS)
Ravindra
R
R
Department of Cell Biology, UMDNJ-School of Osteopathic Medicine, Stratford 08084.
Caro
J F
JF
D000328:Adult; D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000378:metabolism*; D005934:Glucagon / Q000494:pharmacology; D006153:Guanosine Diphosphate / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000378:metabolism; D006165:Guanylyl Imidodiphosphate / Q000494:pharmacology; D006801:Humans; D007328:Insulin / Q000494:pharmacology*; D008099:Liver / Q000378:metabolism*; D008297:Male; D051381:Rats; D017207:Rats, Sprague-Dawley
D007328:Insulin; D006153:Guanosine Diphosphate; D006165:Guanylyl Imidodiphosphate; D006160:Guanosine Triphosphate; D005934:Glucagon; D019204:GTP-Binding Proteins