Solubilization and reconstitution of renal vasopressin receptors

Renal vasopressin receptors (V2) mediating antidiuresis are coupled to adenylate cyclase. To determine the molecular properties of these vasopressin receptors, it is necessary to solubilize the receptors from the membranes. Solubilization of vasopressin receptors in the non-liganded state was shown to abolish hormone recognition. To preserve ligand binding capacity they have developed reconstitution procedures for the renal vasopressin receptors. The pig kidney membranes were solubilized using a zwitterionic detergent, egg lysolecithin and then reconstituted into phospholipid vesicles. Specific binding of [3H] lysine vasopressin [[3H]LVP] to these solubilized reconstituted fractions was fast, saturable and increased linearly with protein concentration. Scatchard analysis of [3H]LVP binding indicated the presence of single class of binding sites with an equilibrium dissociation constant of 2.3 nM. In competition binding experiments, the solubilized receptors displayed the same pharmacological profile as was observed with membrane V2 receptors