title: Development of a cysteine-deprived and C-terminally truncated GLP-1 receptor.
abstract: The glucagon-like peptide-1 receptor (GLP-1R) belongs to family B of the G-protein coupled receptors (GPCRs), and has become a promising target for the treatment of type 2 diabetes. Here we describe the development and characterization of a fully functional cysteine-deprived and C-terminally truncated GLP-1R. Single cysteines were initially substituted with alanine, and functionally redundant cysteines were subsequently changed simultaneously. Our results indicate that Cys(174), Cys(226), Cys(296) and Cys(403) are important for the GLP-1-mediated response, whereas Cys(236), Cys(329), Cys(341), Cys(347), Cys(438), Cys(458) and Cys(462) are not. Extensive deletions were made in the C-terminal tail of GLP-1R in order to determine the limit for truncation. As for other family B GPCRs, we observed a direct correlation between the length of the C-terminal tail and specific binding of (125)I-GLP-1, indicating that the membrane proximal part of the C-terminal is involved in receptor expression at the cell surface. The results show that seven cysteines and more than half of the C-terminal tail can be removed from GLP-1R without compromising GLP-1 binding or function.
authors.affiliation: Department of Incretin Biology, Novo Nordisk, DK-2820 Gentofte, Denmark; Department of Chemistry, MEMPHYS - Center for Biomembrane Physics, Technical University of Denmark, 2800 Kgs. Lyngby, Denmark.
authors.forename: Christina Rye
authors.initials: CR
authors.lastname: Underwood
authors.forename: Lotte Bjerre
authors.initials: LB
authors.lastname: Knudsen
authors.forename: Patrick W
authors.initials: PW
authors.lastname: Garibay
authors.forename: Günther H
authors.initials: GH
authors.lastname: Peters
authors.forename: Steffen
authors.initials: S
authors.lastname: Reedtz-Runge
mesh_terms: D000595:Amino Acid Sequence; D003545:Cysteine / Q000737:chemistry*; D000067757:Glucagon-Like Peptide-1 Receptor; D057809:HEK293 Cells; D006801:Humans; D008969:Molecular Sequence Data; D018027:Receptors, Glucagon / Q000737:chemistry*; D017386:Sequence Homology, Amino Acid
chemical_list: C000593833:GLP1R protein, human; D000067757:Glucagon-Like Peptide-1 Receptor; D018027:Receptors, Glucagon; D003545:Cysteine
keywords: Agonist; CLR; CRFR1; Cysteine; ECL; FRET; G-protein coupled receptor; GCGR; GIP-R; GLP-1; GLP-1R; GPCR; Glucagon-like peptide-1; ICL; JNK1; Mutagenesis; PAC1-R; PTH-1R; PTHrP; TM; VIP; c-Jun N-terminal Kinase 1; calcitonin receptor-like receptor; extracellular loop; fluorescence resonance energy transfer; glucagon receptor; glucagon-like peptide-1; glucagon-like peptide-1 receptor; glucose-dependent insulinotropic polypeptide receptor; intracellular loop; parathyroid hormone related peptide; transmembrane; type-1 corticotrophin-releasing factor receptor; type-1 parathyroid hormone receptor; type-1 pituitary adenylate cyclase-activating polypeptide receptor; vasoactive intestinal polypeptide; β(1/2)-adreneric receptor; β(1/2)AR