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.
D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology*; D007457:Iodine Radioisotopes; D008970:Molecular Weight; D010920:Placenta / Q000032:analysis; D011247:Pregnancy; D011972:Receptor, Insulin / Q000302:isolation & purification*; D012685:Sepharose
D007093:Imidazoles; D007457:Iodine Radioisotopes; C006900:N,N-carbonyldiimidazole; D012685:Sepharose; D011972:Receptor, Insulin