Insulin Receptor Purification
Early work on the purification of the insulin receptor by affinity chromatographic techniques (Cuatrecasas, 1972a) literally set the stage for the small-and large-scale purification of most plasma membrane receptors for hormones and other biologically active agents. Since the early 1970s, little has changed in the basic approach to the isolation of the insulin receptor. The affinity chromatographic isolation protocol played a major role in isolating sufficient amounts of receptor for the microsequence determination, which ultimately, via cloning of the receptor cDNA, led to the elucidation of the complete sequence of the receptor (Ebina et al., 1985; Ullrich et al., 1985). Important modifications of the original isolation protocol have been introduced over the years, including the use of wheat germ agglutinin affinity chromatography (e.g., see Cuatrecasas and Tell, 1973; Harrison and Itin, 1980) and the development of an improved buffer system for the elution of the receptor from the insulin-agarose affinity gel (Fujita-Yamaguchi et al., 1983). The use of wheat germ agglutinin proved superior than the use of concanavalin A; and the omission of urea from the insulin-agarose elution buffer improved the recovery of ligand binding activity and tyrosine kinase activity in the isolated receptor. Apart from insulin-and lectin-agarose affinity chromatography, antireceptor antibodies or anti-insulin antibodies have also proved of use for the affinity-purification of the native or crosslink-labeled receptor (Harrison and Itin, 1980; Siegel et a1.,1981; Heinrich et al., 1980; Armstrong et al., 1982).
KeywordsInsulin Receptor Insulin Binding Ligand Binding Activity Insulin Recep Antireceptor Antibody
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