The catecholamine binding site of the β-adrenergic receptor is formed by juxtaposed membrane-spanning domains

The catecholamine binding domain of the turkey erythrocyte β-adrenergic receptor was mapped by determining the sites of covalent labeling of the purified receptor by two β-adrenergic photoaffinity reagents, [¹²⁵I]iodocyanopindolol-diazirine (ICYP-da) and [¹²⁵I]iodoazidobenzylpindolol (IABP). Both labels were incorporated at two separate sites. By sequencing a labeled peptide, one site of labeling was found to lie at Trp³³⁰ in the extracellular half of the seventh membrane span. This position is homologous to the retinal attachment site in rhodopsin. The second labeled site was isolated on an 8000-Da peptide and immunoprecipitated using sequence-directed antibodies. This site lies in membrane spans 3-5. Labeling of the two sites was equal using ICYP-da and 3-10-fold greater in the span 7 site using IABP. These data indicate that the catecholamine binding site is formed from the juxtaposition of span 7 and spans 3-5 in a tertiary structure probably similar to that of rhodopsin.