11 research outputs found
<新刊紹介>パリ法科大學教授ノガロー原著小樽高等商業學校教授手塚嘉郎著「國際貿易に於ける貨幣の職分と貨幣數量説」
Glucagon-like peptide-1 (GLP-1) activates the GLP-1 receptor (GLP-1R), which belongs to family B of the G-protein-coupled receptors. We previously identified a selective small molecule ligand, compound 2, that acted as a full agonist and allosteric modulator of GLP-1R. In this study, the structurally related small molecule, compound 3, stimulated cAMP production from GLP-1R, but not from the homologous glucagon receptor (GluR). The receptor selectivity encouraged a chimeric receptor approach to identify domains important for compound 3-mediated activation of GLP-1R. A subsegment of the GLP-1R transmembrane domain containing TM2 to TM5 was sufficient to transfer compound 3 responsiveness to GluR. Therefore, divergent residues in this subsegment of GLP-1R and GluR are responsible for the receptor selectivity of compound 3. Functional analyses of other chimeric receptors suggested that the existence of a helix-helix interface between TM1 and TM7 is important for the compound 3 response. Furthermore, site-directed mutagenesis revealed that a Phe195-Leu substitution in TM2 and a Thr391-Ala substitution in TM7 increased and decreased the efficacy of compound 3 without disturbing the potency or efficacy of GLP-1. Collectively, differential effects of receptor mutations suggest that TM2 and/or TM7 are important for compound 3-mediated activation of GLP-1R.</jats:p
α‑Helix or β‑Turn? An Investigation into N‑Terminally Constrained Analogues of Glucagon-like Peptide 1 (GLP-1) and Exendin‑4
Peptide agonists
acting on the glucagon-like peptide 1 receptor
(GLP-1R) promote glucose-dependent insulin release and therefore represent
important therapeutic agents for type 2 diabetes (T2D). Previous data
indicated that an N-terminal type II β-turn motif might be an
important feature for agonists acting on the GLP-1R. In contrast,
recent publications reporting the structure of the full-length GLP-1R
have shown the N-terminus of receptor-bound agonists in an α-helical
conformation. To reconcile these conflicting results, we prepared
N-terminally constrained analogues of glucagon-like peptide 1 (GLP-1)
and exendin-4 and evaluated their receptor affinity and functionality <i>in vitro</i>; we then examined their crystal structures in complex
with the extracellular domain of the GLP-1R and used molecular modeling
and molecular dynamics simulations for further investigations. We
report that the peptides’ N-termini in all determined crystal
structures adopted a type II β-turn conformation, but <i>in vitro</i> potency varied several thousand-fold across the
series. Potency correlated better with α-helicity in our computational
model, although we have found that the energy barrier between the
two mentioned conformations is low in our most potent analogues and
the flexibility of the N-terminus is highlighted by the dynamics simulations
Conformational states of the full-length glucagon receptor
Class B G protein-coupled receptors are composed of an extracellular domain (ECD) and a seven-transmembrane (7TM) domain, and their signalling is regulated by peptide hormones. Using a hybrid structural biology approach together with the ECD and 7TM domain crystal structures of the glucagon receptor (GCGR), we examine the relationship between full-length receptor conformation and peptide ligand binding. Molecular dynamics (MD) and disulfide crosslinking studies suggest that apo-GCGR can adopt both an open and closed conformation associated with extensive contacts between the ECD and 7TM domain. The electron microscopy (EM) map of the full-length GCGR shows how a monoclonal antibody stabilizes the ECD and 7TM domain in an elongated conformation. Hydrogen/deuterium exchange (HDX) studies and MD simulations indicate that an open conformation is also stabilized by peptide ligand binding. The combined studies reveal the open/closed states of GCGR and suggest that glucagon binds to GCGR by a conformational selection mechanism
Discovery of the Once-Weekly Glucagon-Like Peptide‑1 (GLP-1) Analogue Semaglutide
Liraglutide
is an acylated glucagon-like peptide-1 (GLP-1) analogue that binds
to serum albumin <i>in vivo</i> and is approved for once-daily treatment of diabetes as well as obesity. The aim of the present
studies was to design a once weekly GLP-1 analogue by increasing albumin
affinity and secure full stability against metabolic degradation.
The fatty acid moiety and the linking chemistry to GLP-1 were the
key features to secure high albumin affinity and GLP-1 receptor (GLP-1R)
potency and in obtaining a prolonged exposure and action of the GLP-1
analogue. Semaglutide was selected as the optimal once weekly candidate.
Semaglutide has two amino acid substitutions compared to human GLP-1
(Aib<sup>8</sup>, Arg<sup>34</sup>) and is derivatized at lysine 26.
The GLP-1R affinity of semaglutide (0.38 ± 0.06 nM) was three-fold
decreased compared to liraglutide, whereas the albumin affinity was
increased. The plasma half-life was 46.1 h in mini-pigs following i.v.
administration, and semaglutide has an MRT of 63.6 h after s.c. dosing
to mini-pigs. Semaglutide is currently in phase 3 clinical testing
Structure of the full-length glucagon class B G-protein-coupled receptor
The human glucagon receptor, GCGR, belongs to the class B G-protein-coupled receptor family and plays a key role in glucose homeostasis and the pathophysiology of type 2 diabetes. Here we report the 3.0 Å crystal structure of full-length GCGR containing both the extracellular domain and transmembrane domain in an inactive conformation. The two domains are connected by a 12-residue segment termed the stalk, which adopts a β-strand conformation, instead of forming an α-helix as observed in the previously solved structure of the GCGR transmembrane domain. The first extracellular loop exhibits a β-hairpin conformation and interacts with the stalk to form a compact β-sheet structure. Hydrogen-deuterium exchange, disulfide crosslinking and molecular dynamics studies suggest that the stalk and the first extracellular loop have critical roles in modulating peptide ligand binding and receptor activation. These insights into the full-length GCGR structure deepen our understanding of the signalling mechanisms of class B G-protein-coupled receptors