Structural insights into ligand efficacy and activation of the glucagon receptor

Hilger, Daniel; Kumar, Kaavya Krishna; Hu, Hongli; Pedersen, Mogens Theisen; Giehm, Lise; Mathiesen, Jesper Mosolff; Skiniotis, Georgios; Kobilka, Brian K.

doi:10.1101/660837

Cited by 5 publications

(3 citation statements)

References 45 publications

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“…In the structures of the glucagon receptor family members, GLP1R 13,14 and glucagon receptor, GCGR 23,24 , the peptide ligands adopt continuous α-helices and their ECDs cover the ligands to facilitate the interactions with the TMDs, thus playing an indispensable role in receptor activation. Although PACAP also adopts a continuous α-helix, the PAC1R-ECD has no functional role in receptor activation, because PACAP can interact with the TMD without the aid of the ECD.…”

Section: Discussionmentioning

confidence: 99%

Cryo-EM structure of the human PAC1 receptor coupled to an engineered heterotrimeric G protein

Kobayashi

Shihoya

Nishizawa

et al. 2020

Nat Struct Mol Biol

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Pituitary adenylate cyclase-activating polypeptide (PACAP) is a pleiotropic neuropeptide hormone functioning in the central nervous system and peripheral tissues.The PACAP receptor PAC1R, which belongs to the class B G-protein-coupled receptors (GPCRs), is a drug target for mental disorders and dry eye syndrome. Here we present a cryo-electron microscopy structure of human PAC1R bound to PACAP and an engineered Gs heterotrimer. The structure revealed that TM1 plays an essential role in PACAP recognition. The ECD (extracellular domain) of PAC1R tilts by ~40° as compared to that of the glucagon-like peptide-1 receptor (GLP1R), and thus does not cover the peptide ligand. A functional analysis demonstrated that the PAC1R-ECD functions as an affinity trap and is not required for receptor activation, whereas the GLP1R-ECD plays an indispensable role in receptor activation, illuminating the functional diversity of the ECDs in the class B GPCRs. Our structural information will facilitate the design and improvement of better PAC1R agonists for clinical applications. Main textthat for VIP 4 , indicating that PAC1R is relatively selective for PACAP.PAC1R belongs to the class B G-protein-coupled receptors (GPCRs), and predominantly activates the adenylyl cyclase stimulatory G protein Gs. PAC1R is widely expressed in the CNS and peripheral tissues 2 . PACAP/PAC1R signaling has been implicated in playing essential roles in several cellular processes, including circadian rhythm regulation, food intake control, glucose metabolism, learning and memory, neuronal ontogenesis, apoptosis, and immune system regulation. Furthermore, perturbations in the PACAP/PAC1R pathway cause abnormal stress responses underlying posttraumatic stress disorder (PTSD) 5 , and thus PAC1R has been studied as a drug target for numerous disorders. PACAP and PAC1R are expressed in lacrimal glands, and induce tear secretion by increasing the aquaporin 5 (AQP5) levels in the plasma membrane 6 . Therefore, PAC1R is also a drug target for dry eye syndrome.However, the design of small molecule agonists for PAC1R has not yet been achieved, limiting the clinical applications targeting PAC1R.PAC1R comprises two distinct domains: an N-terminal extracellular domain (ECD) and a transmembrane domain (TMD), as in the other class B GPCRs. A two-step/two-domain model has been proposed for ligand binding and receptor activation in the class B GPCRs 7 : the ECD is responsible for the initial and high-affinity binding of peptide ligands, and the TMD plays a key role in both ligand binding and receptor activation. A previous study suggested that PAC1R follows this model, and the PAC1R-ECD is not required for receptor activation 8 . However, in glucagon-like peptide 1 receptor (GLP1R), the ECD also plays an indispensable role in receptor activation, suggesting the divergent role of the ECD in the activation of class B GPCRs. Although the crystal structure of the PAC1R-ECD was determined in a ligand-free conformation 9 , little is known about the mechanism of the ligand recogni...

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Section: Discussionmentioning

confidence: 99%

Cryo-EM structure of the human PAC1 receptor coupled to an engineered heterotrimeric G protein

Kobayashi

Shihoya

Nishizawa

et al. 2020

Nat Struct Mol Biol

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“…Surface expression levels of wild-type and mutant GABA B receptors were determined using a direct enzyme-linked immunosorbent assay (ELISA) against the N-terminal GABA B1b FLAG tag and the N-terminal GABA B2 HA-tag, as described 54 . Transfected cells were seeded in white Poly-D-Lysine-coated 96-well plates.…”

Section: Methodsmentioning

confidence: 99%

Structures of metabotropic GABAB receptor

Papasergi-Scott

Robertson

Seven

et al. 2020

Nature

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GABA (γ-aminobutyric acid) stimulation of the metabotropic GABA B receptor results in prolonged inhibition of neurotransmission that is central to brain physiology 1 . GABA B belongs to the Family C of G protein-coupled receptors (GPCRs), which operate as dimers to relay synaptic neurotransmitter signals into a cellular response through the binding and activation of heterotrimeric G proteins 2 , 3 . GABA B , however, is unique in its function as an obligate heterodimer in which agonist binding and G protein activation take place on distinct subunits 4 , 5 . Here we show structures of heterodimeric and homodimeric full-length GABA B receptors. Complemented by cellular signaling assays and atomistic simulations, the structures reveal an essential role for the GABA B extracellular loop 2 (ECL2) in relaying structural transitions by ordering the linker connecting the extracellular ligand-binding domain to the transmembrane region. Furthermore, the ECL2 of both GABA B subunits caps and interacts with the hydrophilic head of a phospholipid occupying the extracellular half of the transmembrane domain, thereby providing a potentially crucial link between ligand binding and the receptor core that engages G protein. These results provide a starting framework to decipher mechanistic modes of signal transduction mediated by GABA B dimers and have important implications for rational drug design targeting these receptors.

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“…Surface expression levels of wild-type and mutant GABAB receptors were determined using a direct enzyme-linked immunosorbent assay (ELISA) against the N-terminal GABAB1b FLAG tag and the Nterminal GABAB2 HA-tag, as described 57 . Transfected cells were seeded in white Poly-D-Lysine-coated 96-well plates.…”

Section: Cell Surface Elisa Assaymentioning

confidence: 99%

Structures of metabotropic GABA_Breceptor

Papasergi-Scott

Robertson

Seven

et al. 2020

Preprint

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GABA (γ-aminobutyric acid) stimulation of the metabotropic GABAB receptor results in prolonged inhibition of neurotransmission that is central to brain physiology. GABAB belongs to the Family C of G protein-coupled receptors (GPCRs), which operate as dimers to relay synaptic neurotransmitter signals into a cellular response through the binding and activation of heterotrimeric G proteins. GABAB, however, is unique in its function as an obligate heterodimer in which agonist binding and G protein activation take place on distinct subunits. Here we show structures of heterodimeric and homodimeric full-length GABAB receptors. Complemented by cellular signaling assays and atomistic simulations, the structures reveal an essential role for the GABAB extracellular loop 2 (ECL2) in relaying structural transitions by ordering the linker connecting the extracellular ligand-binding domain to the transmembrane region. Furthermore, the ECL2 of both GABAB subunits caps and interacts with the hydrophilic head of a phospholipid occupying the extracellular half of the transmembrane domain, thereby providing a potentially crucial link between ligand binding and the receptor core that engages G protein. These results provide a starting framework to decipher mechanistic modes of signal transduction mediated by GABAB dimers and have important implications for rational drug design targeting these receptors.

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Structural insights into ligand efficacy and activation of the glucagon receptor

Abstract: 16The glucagon receptor family comprises Class B G protein-coupled receptors (GPCRs) that 17 play a crucial role in regulating blood sugar levels. Receptors of this family represent important 18 therapeutic targets for the treatment of diabetes and obesity.

Cited by 5 publications

References 45 publications

Cryo-EM structure of the human PAC1 receptor coupled to an engineered heterotrimeric G protein

Cryo-EM structure of the human PAC1 receptor coupled to an engineered heterotrimeric G protein

Structures of metabotropic GABAB receptor

Structures of metabotropic GABA_Breceptor

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Structural insights into ligand efficacy and activation of the glucagon receptor

Abstract: 16The glucagon receptor family comprises Class B G protein-coupled receptors (GPCRs) that 17 play a crucial role in regulating blood sugar levels. Receptors of this family represent important 18 therapeutic targets for the treatment of diabetes and obesity.

Cited by 5 publications

References 45 publications

Cryo-EM structure of the human PAC1 receptor coupled to an engineered heterotrimeric G protein

Cryo-EM structure of the human PAC1 receptor coupled to an engineered heterotrimeric G protein

Structures of metabotropic GABAB receptor

Structures of metabotropic GABABreceptor

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Product

Resources

About

Structures of metabotropic GABA_Breceptor