A unique hormonal recognition feature of the human glucagon-like peptide-2 receptor

Abstract

Glucagon-like peptides (GLP-1 and GLP-2) are two proglucagon-derived intestinal hormones that mediate distinct physiological functions through two related receptors (GLP-1R and GLP-2R) which are important drug targets for metabolic disorders and Crohn’s disease, respectively. Despite great progress in GLP-1R structure determination, our understanding on the differences of peptide binding and signal transduction between these two receptors remains elusive. Here we report the electron microscopy structure of the human GLP-2R in complex with GLP-2 and a Gs heterotrimer. To accommodate GLP-2 rather than GLP-1, GLP-2R fine-tunes the conformations of the extracellular parts of transmembrane helices (TMs) 1, 5, 7 and extracellular loop 1 (ECL1). In contrast to GLP-1, the N-terminal histidine of GLP-2 penetrates into the receptor core with a unique orientation. The middle region of GLP-2 engages with TM1 and TM7 more extensively than with ECL2, and the GLP-2 C-terminus closely attaches to ECL1, which is the most protruded among 9 class B G protein-coupled receptors (GPCRs). Functional studies revealed that the above three segments of GLP-2 are essential for GLP-2 recognition and receptor activation, especially the middle region. These results provide new insights into the molecular basis of ligand specificity in class B GPCRs and may facilitate the development of more specific therapeutics.

Structural insights into hormone recognition by the human glucose-dependent insulinotropic polypeptide receptor

“…3). In general, the GIPR–G s complex shows a similar receptor–G protein interface as other reported class B1 receptor structures such as GLP-1R ( 27 ), GLP-2R ( 12 ), GCGR ( 14 ), PTH1R ( 16 ), SCTR (secretin receptor) ( 28 ) and GHRHR ( 11 ), suggesting a common G protein signaling mechanism (Fig. 3A).…”

“…To prepare a high quality human GIPR–G s complex, we overcame several technical obstacles to enhance the expression level and protein stability by adding a double tag of maltose binding protein at the C terminus and a BRIL fusion protein at the N terminus (Fig. S1A), as well as employing the NanoBiT tethering strategy ( 10–12 ) (Fig. S1A, B).…”

Evolving cryo-EM structural approaches for GPCR drug discovery

“…Our application of 3D variance analysis to recently solved structures of class B GPCRs, including adenomedullin-1 (AM1) and AM2 receptors 19 , the secretin receptor 20 and GLP-1R bound to peptide and small molecule agonists 14 has revealed common rocking and rotational motions of the G protein relative to the receptors. This is consistent with transient formation and breakage of interactions that are likely to be important in receptor-mediated G protein recruitment and activation, and low resolution for parts of ICL3 and the extension of H8 for class B GPCRs 3,8,9,15,19–28 are consistent with transient interactions between these domains and the G protein. ICL2 exhibited densities consistent with 2 metastable conformations that exhibited differences in the consensus maps solved in the presence and absence of Nb35, and where higher resolution was apparent for the complex without Nb35.…”

“…A key conformational change required nucleotide exchange is outward movement of the AHD of Gs, relative to the ras-like domain. The AHD of Gαs in the nucleotide-free state is highly mobile, thereby occupying different positions around the ras-like domain 30 , and it is poorly resolved in most consensus cryo-EM maps of solved class B GPCR-G protein complexes 3,8,9,15,[19][20][21][22][23][24][25][26][27][28] . When bound with Nb35, 3D multivariate analysis of PF 06882961-GLP-1R-DNGs complex revealed translational motion between open ("up" position) and more closed ("down" position) conformations of the AHD.…”

“…A key tool for stabilisation of GPCR-Gs complexes has been nanobody 35 (Nb35) that binds across the interface of Gs and G subunits, which was first applied to the 2-adrenergic receptor-Gs complex and enabled the first ternary GPCR complex structure to be determined 6 . Alternate or supplementary technologies have subsequently been introduced, including mutant G proteins 7 , nanoBit tethering 8,9 and mini-G proteins that reduce or eliminate guanine nucleotide binding and/or stabilise the interaction between heterotrimeric subunits [10][11][12] . In practice, these have primarily been used in combination with Nb35 for Gs protein complexes to provide enhanced complex stability.…”