Scheel A A, Pelham H R
Medical Research Council Laboratory of Molecular Biology, Cambridge, United Kingdom.
J Biol Chem. 1998 Jan 23;273(4):2467-72. doi: 10.1074/jbc.273.4.2467.
Retention of soluble proteins in the endoplasmic reticulum is dependent on their interaction with the KDEL (Lys-Asp-Glu-Leu) receptor in the Golgi apparatus and their subsequent retrieval back to the endoplasmic reticulum. We have studied the three-dimensional organization of the human KDEL receptor using site-directed mutagenesis and sulfhydryl-specific labeling. We have identified four amino acid residues, Arg-5, Asp-50, Tyr-162, and Asn-165, which we suggest participate in the formation of the ligand binding pocket. Arg-5 and Asp-50 are shown to be located on the lumenal side of the membrane and are inaccessible from the cytoplasm. In addition, our results strongly support a topology of the KDEL receptor similar to the family of G-protein-coupled receptors with seven transmembrane domains. Furthermore, Asp-50 plays a crucial role in the binding of His/Lys-Asp-Glu-Leu ligands, but is not required for Asp-Asp-Glu-Leu binding, suggesting that this residue forms an ion pair with the positively charged amino acid residue positioned 4 residues from the carboxyl terminus of the ligand.
可溶性蛋白质在内质网中的保留取决于它们与高尔基体中KDEL（赖氨酸 - 天冬氨酸 - 谷氨酸 - 亮氨酸）受体的相互作用以及随后返回内质网的过程。我们使用定点诱变和巯基特异性标记研究了人KDEL受体的三维结构。我们确定了四个氨基酸残基，即精氨酸 - 5、天冬氨酸 - 50、酪氨酸 - 162和天冬酰胺 - 165，我们认为它们参与配体结合口袋的形成。精氨酸 - 5和天冬氨酸 - 50位于膜的腔侧，从细胞质无法接近。此外，我们的结果有力地支持了KDEL受体的拓扑结构类似于具有七个跨膜结构域的G蛋白偶联受体家族。此外，天冬氨酸 - 50在组氨酸/赖氨酸 - 天冬氨酸 - 谷氨酸 - 亮氨酸配体的结合中起关键作用，但对于天冬氨酸 - 天冬氨酸 - 谷氨酸 - 亮氨酸的结合不是必需的，这表明该残基与位于配体羧基末端4个残基处的带正电荷的氨基酸残基形成离子对。
