This chapter discusses the affinity labeling of receptors for steroid and thyroid hormones. Affinity labeling is a technique for the selective, covalent labeling of binding sites in macromolecules. It involves initially the reversible binding of an affinity probe, which is a ligand modified to contain a reactive functional group (attaching function), within the binding site of the macromolecule, followed then by the covalent attachment of the affinity probe to the molecular constituents within or near the binding site. The selectivity of the labeling process derives from the selective interaction of the affinity probe with the binding site. The labeling of receptors for hormones generally requires that a very small quantity of protein, generally present as a very minor component in a heterogeneous binding preparation, undergo covalent interaction with the affinity probe. For this to be done successfully, or at best optimally, the labeling process must be both efficient—that is, as much as possible (ideally all) of the receptor should become labeled—and selective—that is, as little as possible of the other non-receptor proteins should become labeled.
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