PMID:22147709

Citation	Koole, C, Wootten, D, Simms, J, Savage, EE, Miller, LJ, Christopoulos, A and Sexton, PM (2012) Second extracellular loop of human glucagon-like peptide-1 receptor (GLP-1R) differentially regulates orthosteric but not allosteric agonist binding and function. J. Biol. Chem. 287:3659-73
Abstract	The glucagon-like peptide-1 receptor (GLP-1R) is a prototypical family B G protein-coupled receptor that exhibits physiologically important pleiotropic coupling and ligand-dependent signal bias. In our accompanying article (Koole, C., Wootten, D., Simms, J., Miller, L. J., Christopoulos, A., and Sexton, P. M. (2012) J. Biol. Chem. 287, 3642-3658), we demonstrate, through alanine-scanning mutagenesis, a key role for extracellular loop (ECL) 2 of the receptor in propagating activation transition mediated by GLP-1 peptides that occurs in a peptide- and pathway-dependent manner for cAMP formation, intracellular (Ca(2+)(i)) mobilization, and phosphorylation of extracellular signal-regulated kinases 1 and 2 (pERK1/2). In this study, we examine the effect of ECL2 mutations on the binding and signaling of the peptide mimetics, exendin-4 and oxyntomodulin, as well as small molecule allosteric agonist 6,7-dichloro-2-methylsulfonyl-3-tert-butylaminoquinoxaline (compound 2). Lys-288, Cys-296, Trp-297, and Asn-300 were globally important for peptide signaling and also had critical roles in governing signal bias of the receptor. Peptide-specific effects on relative efficacy and signal bias were most commonly observed for residues 301-305, although R299A mutation also caused significantly different effects for individual peptides. Met-303 was more important for exendin-4 and oxyntomodulin action than those of GLP-1 peptides. Globally, ECL2 mutation was more detrimental to exendin-4-mediated Ca(2+)i release than GLP-1(7-36)-NH(2), providing additional evidence for subtle differences in receptor activation by these two peptides. Unlike peptide activation of the GLP-1R, ECL2 mutations had only limited impact on compound 2 mediated cAMP and pERK responses, consistent with this ligand having a distinct mechanism for receptor activation. These data suggest a critical role of ECL2 of the GLP-1R in the activation transition of the receptor by peptide agonists.
Links	PubMed PMC3281730 Online version:10.1074/jbc.M111.309369
Keywords	Allosteric Regulation; Amino Acid Substitution; Biomimetic Materials/chemistry; Biomimetic Materials/pharmacology; Cell Line; Humans; MAP Kinase Signaling System/drug effects; MAP Kinase Signaling System/physiology; Mitogen-Activated Protein Kinase 1/chemistry; Mitogen-Activated Protein Kinase 1/genetics; Mitogen-Activated Protein Kinase 1/metabolism; Mitogen-Activated Protein Kinase 3/chemistry; Mitogen-Activated Protein Kinase 3/genetics; Mitogen-Activated Protein Kinase 3/metabolism; Mutation, Missense; Peptides/chemistry; Peptides/pharmacology; Protein Structure, Secondary; Protein Structure, Tertiary; Receptors, Glucagon/chemistry; Receptors, Glucagon/genetics; Receptors, Glucagon/metabolism
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
HUMAN:GLP1R		GO:0008528: G-protein coupled peptide receptor activity	ECO:0000314:		F		Shown through an experiment causing mutations, the ECL2 of the GLP-1R plays a large role in GLP-1 peptide-mediated selective signaling and receptor activation. Detailed on pages 3-4	complete CACAO 3567

References

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