In order to fully understand how receptors function, it is necessary to purify them and characterize their structures and biochemical properties in detail. Determining their molecular structures with and without a bound ligand, for instance, can elucidate the conformational changes that occur upon ligand binding that activate downstream signal transduction proteins. But separation of cell-surface receptors from other cellular proteins is very challenging. A “typical” mammalian cell has 1000 to 50,000 copies of a single type of cell-surface receptor. This may seem like a large number, but when you consider that this same cell contains ~10¹⁰ total protein molecules, with ~10⁶ of them in the plasma membrane alone, you realize that these receptors constitute only 0.1 to 5 percent of plasma-membrane proteins. This low abundance complicates the isolation and purification of cell-surface receptors. Purification of these integral membrane proteins is also difficult because the membrane must first be solubilized with a non-ionic detergent (see Figure 7-23) under conditions in which the three-dimensional structure of the receptor and its ability to bind ligand are maintained. Then the receptors can be separated from other cellular molecules.

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Recombinant DNA techniques can often be used to generate cells that express large amounts of receptor proteins. But even when such techniques are used, special techniques are necessary to isolate the receptors from other membrane proteins. One technique often used in purifying cell-surface receptors that retain their ligand-binding ability when they are solubilized by detergents is a type of affinity chromatography (see Figure 3-40c). An antibody that recognizes either the receptor or a ligand for the receptor is chemically linked to the beads used to form a column. A crude, detergent-solubilized preparation of membrane proteins is then passed through the column. Only the receptor protein, together with other proteins tightly bound to it, will specifically stick to the column; other proteins are washed away. Once the other proteins are removed, the receptors can be released (“eluted”) from the column either by passage of an excess of the soluble ligand through the column (ligand affinity chromatography) or by using chemical conditions (e.g., changes in pH) to release the receptor from the antibody (antibody affinity chromatography). In some cases, a receptor can be purified as much as 100,000-fold in a single affinity-chromatographic step.