Identification of amino acids in the binding pocket of the human KDEL receptor.
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.
Scheel
A A
AA
Medical Research Council Laboratory of Molecular Biology, Cambridge, United Kingdom.
Pelham
H R
HR
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000596:Amino Acids / Q000737:chemistry* / Q000378:metabolism; D001665:Binding Sites; D003545:Cysteine / Q000737:chemistry; D005033:Ethylmaleimide / Q000494:pharmacology; D006801:Humans; D008024:Ligands; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017433:Protein Structure, Secondary; D018000:Receptors, Peptide / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D013439:Sulfhydryl Reagents / Q000494:pharmacology
D000596:Amino Acids; C064459:KDEL receptor; D008024:Ligands; D018000:Receptors, Peptide; D013439:Sulfhydryl Reagents; D003545:Cysteine; D005033:Ethylmaleimide