Perlman J H, Colson A O, Wang W, Bence K, Osman R, Gershengorn M C
Division of Molecular Medicine, Department of Medicine, Cornell University Medical College and The New York Hospital, New York, New York 10021, USA.
J Biol Chem. 1997 May 2;272(18):11937-42. doi: 10.1074/jbc.272.18.11937.
The roles of conserved residues in transmembrane helices (TMs) of G protein-coupled receptors have not been well established. A computer-generated model of the thyrotropin-releasing hormone receptor (TRH-R) indicated that conserved Asp-71 (TM-2) could interact with conserved asparagines 316 (TM-7) and 43 (TM-1). To test this model, we constructed mutant TRH-Rs containing polar or alanine substitutions of these residues. The maximal activities of N43A and N316A TRH-Rs were diminished, whereas D71A (Perlman, J. H., Nussenzveig, D. R., Osman, R., and Gershengorn, M. C. (1992) J. Biol. Chem. 267, 24413-24417) and N43A/N316A TRH-Rs were inactive. Computer models of D71A and N43A/N316A TRH-Rs show similar changes from native TRH-R in their TM bundle conformations. The inactivity and the similarity of the computer models of D71A and N43A/N316A TRH-Rs are consistent with the idea that Asp-71 bridges Asn-43 and Asn-316 and suggest that activity is critically dependent on these interactions. The conservation of these residues suggests these specific interactions involving TMs 1, 2, and 7 may be structurally important for all members of the rhodopsin/beta-adrenergic receptor subfamily of G protein-coupled receptors.
G蛋白偶联受体跨膜螺旋（TMs）中保守残基的作用尚未完全明确。促甲状腺激素释放激素受体（TRH-R）的计算机生成模型表明，保守的天冬氨酸-71（TM-2）可与保守的天冬酰胺316（TM-7）和43（TM-1）相互作用。为验证该模型，我们构建了这些残基被极性氨基酸取代或丙氨酸取代的突变型TRH-R。N43A和N316A突变型TRH-R的最大活性降低，而D71A（佩尔曼，J.H.，努森兹韦格，D.R.，奥斯曼，R.，和格申戈恩，M.C.（1992年）《生物化学杂志》267卷，24413 - 24417页）以及N43A/N316A突变型TRH-R无活性。D71A和N43A/N316A突变型TRH-R的计算机模型显示，其TM束构象与天然TRH-R相比有类似变化。D71A和N43A/N316A突变型TRH-R的无活性以及计算机模型的相似性与天冬氨酸-71连接天冬酰胺-43和天冬酰胺-316的观点一致，并表明活性关键依赖于这些相互作用。这些残基的保守性表明，这些涉及TMs 1、2和7的特定相互作用可能对G蛋白偶联受体视紫红质/β-肾上腺素能受体亚家族的所有成员在结构上具有重要意义。
