Identification of amino acids in the binding pocket of the human KDEL receptor

Abstract

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.