Solubilization and separation of the glucagon receptor and adenylate cyclase in guanine nucleotide-sensitive states.
Adenylate cyclase in liver membranes was solubilized with Lubrol PX and partially purified by gel filtration. The partially purified enzyme was susceptible to activation by guanyl-5'-yl imidodiphosphate (Gpp(NH)p). Studies on the binding of [3H]Gpp(NH)p to various fractions eluted from the gels revealed that an upper limit of 1% of the Gpp(NH)p binding sites is associated with adenylate cyclase activity stimulated by the nucleotide. The glucagon receptor, pretagged with 125I-glucagon in the membranes, solubilized with Lubrol PX, and fractionated on the same gel columns, eluted in a peak fraction that overlaps with, but is separate from, adenylate cyclase in its Gpp(NH)p-stimulated form. Addition of GTP to the solubilized glucagon-receptor complex caused complete dissociation of the complex, as has been shown with the membrane-bound form of the complex. Since the GTP-sensitive form of the glucagon receptor complex separates from the Gpp(NH)p-sensitive form of adenylate cyclase, it is concluded that the receptor and the enzyme are separate molecules, each associated with a distinct nucleotide regulatory site or component. These findings are discussed in terms of the possible structure of the hormone-sensitive state of adenylate cyclase.
D000262:Adenylyl Cyclases / Q000302:isolation & purification / Q000378:metabolism; D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D005934:Glucagon / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology; D008099:Liver / Q000378:metabolism; D051381:Rats; D011956:Receptors, Cell Surface / Q000302:isolation & purification / Q000378:metabolism; D011955:Receptors, Drug / Q000302:isolation & purification; D012995:Solubility
D011956:Receptors, Cell Surface; D011955:Receptors, Drug; D006160:Guanosine Triphosphate; D005934:Glucagon; D000262:Adenylyl Cyclases