A Hydrogen-Bonded Polar Network in the Core of the Glucagon-Like Peptide-1 Receptor Is a Fulcrum for Biased Agonism: Lessons from Class B Crystal Structures

The glucagon-like peptide 1 (GLP-1) receptor is a class B G protein-coupled receptor (GPCR) that is a key target for treatments for type II diabetes and obesity. This receptor, like other class B GPCRs, displays biased agonism, though the physiologic significance of this is yet to be elucidated. Previous work has implicated R2.60¹⁹⁰ , N3.43²⁴⁰ , Q7.49³⁹⁴ , and H6.52³⁶³ as key residues involved in peptide-mediated biased agonism, with R2.60¹⁹⁰ , N3.43²⁴⁰ , and Q7.49³⁹⁴ predicted to form a polar interaction network. In this study, we used novel insight gained from recent crystal structures of the transmembrane domains of the glucagon and corticotropin releasing factor 1 (CRF1) receptors to develop improved models of the GLP-1 receptor that predict additional key molecular interactions with these amino acids. We have introduced E6.53³⁶⁴ A, N3.43²⁴⁰ Q, Q7.49⁴⁹³N, and N3.43²⁴⁰ Q/Q7.49 Q/Q7.49⁴⁹³N mutations to probe the role of predicted H-bonding and charge-charge interactions in driving cAMP, calcium, or extracellular signal-regulated kinase (ERK) signaling. A polar interaction between E6.53³⁶⁴ and R2.60190 was predicted to be important for GLP-1- and exendin-4-, but not oxyntomodulin-mediated cAMP formation and also ERK1/2 phosphorylation. In contrast, Q7.49394 , but not R2.60¹⁹⁰ /E6.53³⁶⁴ was critical for calcium mobilization for all three peptides. Mutation of N3.43²⁴⁰ and Q7.49³⁹⁴ had differential effects on individual peptides, providing evidence for molecular differences in activation transition. Collectively, this work expands our understanding of peptide-mediated signaling from the GLP-1 receptor and the key role that the central polar network plays in these events.