Selective solubilization of physalaemin-type substance P binding sites from rat brain membranes by glycodeoxycholate and NaCl

Abstract

Active substance P binding sites were solubilized from rat brain membranes by treatment with 0.125% sodium glycodeoxycholate and 1 M NaCl. About 50% of the binding activity in membrane-bound binding sites was recovered in the solubilized fraction after centrifugation at 105,000 g for 1 h. [ 3H]Substance P absorbed extensively to glass tubes and glass filters, but the absorption was greatly reduced by siliconizing glass tubes and preincubating glass filters in a solution containing poly- d-lysine and bovine serum albumin. [ 3H]Substance P was found to bind the solubilized receptors in a saturable fashion with a B max of 145 fmol/mg protein and a K d of 4.6 nM, and these bindings were completely replaced by low concentrations of unlabeled substance P and physalaemin.