Dynamics of GLP-1R peptide agonist engagement are correlated with kinetics of G protein activation

Abstract

The glucagon-like peptide-1 receptor (GLP-1R) has broad physiological roles and is a validated target for treatment of metabolic disorders. Despite recent advances in GLP-1R structure elucidation, detailed mechanistic understanding of how different peptides generate profound differences in G protein-mediated signalling is still lacking. Here we have combined cryo-electron microscopy, molecular dynamics simulations, receptor mutagenesis and pharmacological assays, to interrogate the mechanism and consequences of GLP-1R binding by four peptide agonists; glucagon-like peptide-1, oxyntomodulin, exendin-4 and exendin-P5. These data revealed that distinctions in peptide N-terminal interactions and dynamics with the GLP-1R transmembrane domain are reciprocally associated with differences in the allosteric coupling to G proteins. In particular, transient interactions with residues at the base of the binding cavity correlate with enhanced kinetics for G protein activation, providing a rationale for differences in G protein-mediated signalling efficacy from distinct agonists.

Structural and functional diversity among agonist-bound states of the GLP-1 receptor

“…However, in addition to being supported by data presented above, our two-state GLP-1R activation hypothesis is consistent with previous studies that support a role for ligand mobility in activation of other GPCRs. [47][48][49] Our findings are distinct from these precedents, however, in suggesting that at least two distinct states of an agonist-receptor complex play important and complementary roles in the signal transduction mechanism.…”

“…The missing stalk region of GLP-1R (residues 129-137) was added to models derived from GLP-1R:Ex4-D-Ala:Gs (conformer 1) using rabbit GLP-1R sequence (PDB: 5VAI) 8 by superposition on the structure of GLP-1R:Ex4:Gs from previous work. 49 The Gs protein was removed, with the exception of Gα helix h5 (residues 370-394), which was retained to keep GLP-1R in fully active conformation during the simulations (in line with our previous work 9 ). The resulting simplified GLP-1R:Ex4-D-Ala:Gα(h5) system was prepared for simulations with the CHARMM36 64 force field using VMD 65 and in-house python HTMD 66 and Tcl (Tool Command Language) scripts.…”

“…Our conclusions are consistent with emerging evidence that conformational mobility in agonist and receptor can be functionally important in signal-transducing states of other GPCRpeptide complexes. [47][48][49] The mode of agonist mobility highlighted in this work may be evolutionarily conserved among peptide agonists of related GPCRs; Gly at the fourth position from the N-terminus is found in glucagon, GLP-2 and several other hormones. 16 Other sites of essential mobility may be present in more distantly related hormones, such as parathyroid hormone.…”

Random glucose GWAS in 493,036 individuals provides insights into diabetes pathophysiology, complications and treatment stratification

“…To gain structural insights into GLP1R variant effects we performed molecular dynamics simulations of the human GLP-1R bound to oxyntomodulin 27 ( Extended Data Tables 1-6 ). A316T has a single amino acid substitution in the core of the receptor transmembrane domain ( Figure 2c ) that leads to an alteration of the hydrogen bond network in close proximity ( Video S1 ).…”

“…The active state structure of GLP-1R in complex with OXM 27 and Gs protein was modelled as previously described 30 and used to simulate the WT GLP-1R and G168S, A316T and R421W. The systems were prepared for molecular dynamics (MD) simulations and equilibrated as reported in 30 .…”