Biology subjects

Sloop, K. W.

Publications and source records attributed to Sloop, K. W..

3 recordsLinked to original sources

bioRxiv2026

Structural dynamics underlying agonist activation of a GLP-1R-Gs precoupled complex

G-protein-coupled receptors (GPCRs) act as allosteric transmembrane signalling machines, generating distinct cellular responses depending on the conformational states induced by ligand binding. The glucagon-like peptide-1 receptor (GLP-1R), a class B GPCR central to insulin secretion and body-weight regulation, is a key therapeutic target for obesity-associated metabolic disease. Here, we used hydrogen-deuterium exchange mass spectrometry to characterize ligand-evoked structural dynamics within a pre-coupled GLP-1R-Gs protein complex. Non-peptide agonists Chu-128 and danuglipron elicited overlapping dynamic perturbation profiles, with distinct drug-specific effects within the transmembrane bundle. In contrast, the natural GLP-1 hormone produced a weaker stabilizing effect on receptor backbone dynamics, while its inactive metabolite exerted opposing localised destabilization. Notably, both peptides uniquely modulated the highly flexible G-protein switch III loop, a key mediator of downstream signalling. These findings pinpoint areas where structural dynamics shape agonist efficacy and facilitate functional dynamics-integrated drug discovery of non-peptide agonists. SignificanceThe development of non-peptide agonists of the glucagon-like peptide-1 receptor (GLP-1R) represents a major advance in metabolic therapeutics, addressing key limitations of current peptide-based incretin therapies while enabling improved control over receptor signalling and pharmacokinetic properties. The recent FDA approval of a first-in-class small-molecule oral GLP-1R agonist, LY3502970, highlights the translational potential of this approach. However, the molecular basis by which distinct ligands modulate GLP-1R conformational dynamics and signalling remains poorly understood. Here we report how non-peptide agonists (Chu-128 and danuglipron) and endogenous GLP-1 peptide and its inactive metabolite shape the structural dynamics of a pre-coupled GLP-1R-Gs complex, revealing patterns linked to receptor activation. Our findings provide insights that guide the rational design of next-generation GLP-1R therapeutics.

biochemistry↗

bioRxiv2024

The physiological impact of an N-terminal Halo-tag on GIPR function in mice

AimsG protein-coupled receptors (GPCRs) modified with self-labelling enzymatic tags (e.g. Halo, SNAP, CLIP) have enabled the in vitro study of receptor expression and trafficking. We developed a GiprHalo/Halo mouse model to gain insights into endogenous glucose-dependent insulinotropic polypeptide (GIP) receptor (GIPR) signalling, an important modulator of glucose and appetite homeostasis. MethodsGiprHalo/Halo mice were generated through ES cell-based gene targeting and physiologically characterised via body weight, oral glucose tolerance test and intraperitoneal glucose tolerance test analysis. Live cell imaging was used to measure cAMP responses in dispersed pancreatic islets from GiprWt/Wt and GiprHalo/Halo littermates in response to GIP. cAMP accumulation and surface receptor expression were assessed in AD293 cells transiently transfected with mouse and human untagged GIPR and Halo-GIPR. Endogenous pancreatic islet expression of Halo-GIPR was determined by immunohistochemistry. ResultsGiprHalo/Halo mice displayed comparable body weights and responses to oral glucose administration as GiprHalo/Wt and GiprWt/Wt littermates. However, GiprHalo/Halo mice only responded to high dose human GIP during intraperitoneal glucose tolerance testing, and cAMP responses to GIP were impaired in GiprHalo/Halo islets. Despite unimpaired surface expression, in vitro cAMP responses were lessened in AD293 cells expressing Halo-GIPR compared to untagged GIPR. Anti-Halo staining in mouse GiprHalo/Wt pancreatic islets displayed detectable signal. ConclusionsWhile the GiprHalo/Halo mouse model could be used to study endogenous GIPR expression, due to reduced responses we advise against its use for endogenous GIPR trafficking and signalling. Our study highlights the importance of carefully phenotyping mouse models in which modifications to endogenous proteins have been introduced.

cell biology↗

bioRxiv2024

In vivo functional profiling and structural characterisation of the human Glp1r A316T variant

Glucagon-like peptide-1 receptor agonists (GLP-1RAs) are effective therapies for type 2 diabetes (T2D) and obesity, yet patient responses are variable. Variation in the human Glp1r gene might be directly linked to therapeutic responses. A naturally occurring missense variant, A316T, protects against T2D and cardiovascular disease. Here, we have generated and characterised a human Glp1r A316T mouse model. Human Glp1rA316T/A316T mice displayed lower fasting blood glucose versus wildtype littermates, even under metabolic stress, and exhibited alterations in islet cytoarchitecture and /{beta}-cell identity under a high-fat, high-sucrose diet. This was however associated with blunted responses to GLP-1RAs in vivo. Further investigations in rodent and human {beta}-cell models demonstrated that human Glp1r A316T exhibits characteristics of constitutive activation but dampened GLP-1RA responses. Results are further supported by cryo-EM analyses and molecular dynamics simulations of GLP-1R A316T structure, collectively demonstrating that the A316T variant governs basal GLP-1R activity and pharmacological responses to GLP-1R-targeting therapies. TeaserThe Glp1r A316T missense variant displays improved glucose tolerance but impaired pharmacological incretin responses in vivo.

cell biology↗