Homogeneous bivalent insulin receptor: purification using insulin coupled to 1,1'-carbonyldiimidazole activated-agarose.

A novel affinity gel, consisting of insulin coupled to 1,1'-carbonyldiimidazole-activated agarose (CDI-agarose), was used to purify insulin receptors from human placenta to homogeneity. This affinity gel is reproducibly prepared and is reported to have a number of advantages over the standard cyanogen bromide activated supports, such as ease and simplicity of coupling and minimal ligand leakage and non-specific binding. Insulin receptors in Triton X-100-solubilized microsomal membranes were purified 2,000-fold by sequential affinity chromatography on wheat germ lectin-agarose and insulin-CDI-activated agarose. They have one of the highest specific insulin-binding capacities (6 nmol/mg protein) reported and can be calculated to have a binding valence of two on the basis that the molecular weight of the oligomeric receptor is 300-350,000.