Wong S K, Slaughter C, Ruoho A E, Ross E M
Department of Pharmacology, Southwestern Graduate School of Biomedical Sciences, University of Texas Health Science Center, Dallas 75235-9041.
J Biol Chem. 1988 Jun 15;263(17):7925-8.
The catecholamine binding domain of the turkey erythrocyte beta-adrenergic receptor was mapped by determining the sites of covalent labeling of the purified receptor by two beta-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.
通过确定两种β-肾上腺素能光亲和试剂[125I]碘氰吲哚洛尔-重氮嗪（ICYP-da）和[125I]碘叠氮苄基吲哚洛尔（IABP）对纯化受体的共价标记位点，绘制了火鸡红细胞β-肾上腺素能受体的儿茶酚胺结合结构域。两种标记都在两个不同的位点掺入。通过对一个标记肽进行测序，发现一个标记位点位于第七个膜跨段细胞外部分的Trp330处。这个位置与视紫红质中的视黄醛附着位点同源。第二个标记位点在一个8000道尔顿的肽上分离出来，并使用序列导向抗体进行免疫沉淀。这个位点位于膜跨段3-5中。使用ICYP-da时，两个位点的标记相等，而使用IABP时，在跨段7位点的标记比其他位点大3至10倍。这些数据表明，儿茶酚胺结合位点是由跨段7与跨段3-5在可能类似于视紫红质的三级结构中并列形成的。
