1,6-α-Aminosuberic acid, 3-(p-azidophenylalanine),8-arginine]vasopressin as a photoaffinity label for renal vasopressin receptors: An evaluation

Abstract

The photoreactive analog of vasopressin [1,6-α-aminosuberic acid, 3-(p-azidophenylalanine),8-arginine]vasopressin was labeled with tritium (specific radioactivity: 39 Ci/mmole). In the absence of UV-light, the tritium-labeled ligand retained a high binding affinity to the vasopressin receptor in plasma membranes from bovine kidney inner medulla (apparent dissociation constant KD: 1.4 × 10−8 M). Renal plasma membranes were irradiated under conditions of a high ratio of specific versus non-specific binding of the reactive ligand. Sodium dodecyl sulfate gel electrophoresis after solubilization and reduction demonstrated the preferential labeling of a polypeptide with an apparent molecular weight of Mr = 32 000. A comparison with stained gels revealed that this protein is a minor constitutent of the bovine kidney membrane. Our results suggest the 32 000-dalton polypeptide is either a component of the vasopressin receptor or that it is located in the immediate vicinity of the vasopressin binding protein.