Properties of rat erythrocyte membrane cytoskeletal structures produced by digitonin extraction: digitonin-insoluble beta-adrenergic receptor, adenylate cyclase, and cholera toxin substrate.
Rat erythrocyte plasma membranes have been extracted exhaustively with digitonin at low temperature, and the residual, detergent-extracted membrane cytoskeletal material is compared to that prepared with Triton X-100 with respect to protein, glycoprotein, phospholipid, and cholesterol content. Digitonin, a weaker detergent than Triton X-100, solubilizes only 26% of the phospholipids and none of the cholesterol. SDS-polyacrylamide gel electrophoresis reveals that differences between the proteins extracted by the two detergents are primarily quantitative. In terms of functional preservation, digitonin retains in the cytoskeleton 28% of the beta-adrenergic receptor binding activity (with the balance accounted for in the supernatant), greater than 90% of the adenylate cyclase and greater than 90% of the 45,000 mol wt polypeptide cholera toxin substrate. The cytoskeletal-associated beat-adrenergic receptor retains binding properties for antagonist and agonist which are identical to those of the native membrane receptor. The digitonin-extracted cytoskeleton containing the beta-adrenergic receptor may provide a useful vehicle for the reconstitution of a hormone-sensitive adenylate cyclase.
D000262:Adenylyl Cyclases / Q000097:blood* / Q000302:isolation & purification; D000818:Animals; D002772:Cholera Toxin / Q000378:metabolism*; D004072:Digitonin; D004910:Erythrocyte Membrane / Q000378:metabolism*; D004912:Erythrocytes / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D008563:Membrane Lipids / Q000097:blood / Q000302:isolation & purification; D051381:Rats; D011919:Rats, Inbred Strains; D011941:Receptors, Adrenergic / Q000378:metabolism*; D011943:Receptors, Adrenergic, beta / Q000302:isolation & purification / Q000378:metabolism*; D012995:Solubility
D008563:Membrane Lipids; D011941:Receptors, Adrenergic; D011943:Receptors, Adrenergic, beta; D002772:Cholera Toxin; D000262:Adenylyl Cyclases; D004072:Digitonin