Evidence for complex formation between GTP binding protein(Gs) and membrane-associated nucleoside diphosphate kinase.
When the Gs in rat liver membranes was prelabeled with [32P]NAD and cholera toxin, solubilized with octylglucoside, and then analyzed by sucrose density gradient centrifugation, it was fractionated into two peaks with approximate molecular sizes of 12-13S and 3-4S. Pretreatment without or with GDP beta S of the labeled membranes resulted in a larger peak in the high molecular weight region, whereas pretreatment with glucagon plus GTP gamma S caused almost equal peaks in both regions. The affinity-purified anti-nucleoside diphosphate (NDP) kinase antibodies only precipitated the Gs in high molecular weight region. Under the same condition, small but significant NDP kinase activity was associated with the high molecular weight Gs region although a large portion of the enzyme activity was recovered in fractions where it alone should appear (6.2S). Both Lubrol-PX and digitonin solubilized the Gs in forms insensitive to immunoprecipitation by anti-NDP kinase antibodies although the latter detergent was able to solubilize the Gs in a high molecular weight form, that is, a ternary glucagon-receptor-G protein complex. These results demonstrate that Gs and membrane-associated NDP kinase may exist in part in a complexed form in membranes. Physiological relevance of the complex formation in membrane signal transduction is discussed.
Kimura
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Department of Biochemistry, Tokyo Metropolitan Institute of Gerontology, Japan.
Shimada
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N
D000246:Adenosine Diphosphate Ribose / Q000378:metabolism; D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D002499:Centrifugation, Density Gradient; D019204:GTP-Binding Proteins / Q000378:metabolism*; D008099:Liver / Q000378:metabolism; D046911:Macromolecular Substances; D009701:Nucleoside-Diphosphate Kinase / Q000378:metabolism*; D010770:Phosphotransferases / Q000378:metabolism*; D011485:Protein Binding; D051381:Rats; D011964:Receptors, Gastrointestinal Hormone / Q000378:metabolism; D018027:Receptors, Glucagon; D012995:Solubility
D046911:Macromolecular Substances; D011964:Receptors, Gastrointestinal Hormone; D018027:Receptors, Glucagon; D000246:Adenosine Diphosphate Ribose; D010770:Phosphotransferases; D009701:Nucleoside-Diphosphate Kinase; D019204:GTP-Binding Proteins; D000262:Adenylyl Cyclases