title: Molecular characterization of the solubilized atrial natriuretic factor receptor from bovine adrenal zona glomerulosa.
abstract: The atrial natriuretic factor (ANF) receptor has been solubilized from bovine adrenal zona glomerulosa membranes with the nonionic detergent octyl-beta-D-glucoside. Mathematical analysis of competition binding curves with solubilized receptor revealed the presence of two classes of binding sites with pK of 10.4 (Kd = 40 pM) and 8.2 (kd = 6000 pM), similar to the native receptor of intact membranes. The hydrodynamic properties of the ANF receptor were determined by prelabeling the membrane receptor with 125I-ANF prior to solubilization. The solubilized 125I-ANF-receptor complex eluted as a major peak with a Stokes radius of 50.8 A from a Superose 6 steric exclusion column. A partial specific volume of 0.770 ml/g and a sedimentation coefficient (S20,w) of 6.34 S were determined by sucrose density gradient centrifugation in H2O and D2O. These data were used to calculate a molecular weight of 158,000 and a frictional ratio of 1.25 for the labeled receptor-detergent complex. The amount of detergent bound to the receptor was estimated to be 0.45 g/g of protein, assuming a partial specific volume of 0.730 ml/g for the protein. Correction for the mass contributed by the bound detergent yielded a molecular weight of 109,000 for the receptor protein. Affinity cross-linking of 125I-ANF to its binding sites in zona glomerulosa membranes and analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography revealed that one single band with apparent Mr 130,000 was specifically labeled. These results indicate that the ANF receptor from bovine adrenal cortex is a membrane protein with a total molecular weight of 110,000-130,000 and suggest that the native protein contains only one single polypeptide chain.
mesh_terms: D000302:Adrenal Cortex / Q000378:metabolism*; D000818:Animals; D009320:Atrial Natriuretic Factor / Q000378:metabolism*; D002417:Cattle; D002462:Cell Membrane / Q000378:metabolism; D007700:Kinetics; D008970:Molecular Weight; D011487:Protein Conformation; D017461:Receptors, Atrial Natriuretic Factor; D011956:Receptors, Cell Surface / Q000302:isolation & purification / Q000378:metabolism*; D012995:Solubility
chemical_list: D011956:Receptors, Cell Surface; D009320:Atrial Natriuretic Factor; D017461:Receptors, Atrial Natriuretic Factor