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- PDB-6vcb: Cryo-EM structure of the Glucagon-like peptide-1 receptor in comp... -

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Basic information

Entry
Database: PDB / ID: 6vcb
TitleCryo-EM structure of the Glucagon-like peptide-1 receptor in complex with G protein, GLP-1 peptide and a positive allosteric modulator
Components
  • (Glucagon-like peptide ...) x 2
  • (Guanine nucleotide-binding protein ...) x 3
  • Nanobody 35
KeywordsSIGNALING PROTEIN/MEMBRANE PROTEIN / G protein-coupled receptor / membrane protein / family B GPCR / diabetes / allosteric modulator / SIGNALING PROTEIN-MEMBRANE PROTEIN complex
Function / homology
Function and homology information


glucagon receptor binding / glucagon-like peptide 1 receptor activity / glucagon receptor activity / protein kinase A signaling / hormone secretion / positive regulation of blood pressure / post-translational protein targeting to membrane, translocation / negative regulation of execution phase of apoptosis / feeding behavior / response to psychosocial stress ...glucagon receptor binding / glucagon-like peptide 1 receptor activity / glucagon receptor activity / protein kinase A signaling / hormone secretion / positive regulation of blood pressure / post-translational protein targeting to membrane, translocation / negative regulation of execution phase of apoptosis / feeding behavior / response to psychosocial stress / regulation of heart contraction / positive regulation of calcium ion import / peptide hormone binding / positive regulation of insulin secretion involved in cellular response to glucose stimulus / PKA activation in glucagon signalling / hair follicle placode formation / Synthesis, secretion, and deacylation of Ghrelin / developmental growth / D1 dopamine receptor binding / intracellular transport / renal water homeostasis / Hedgehog 'off' state / adenylate cyclase-activating adrenergic receptor signaling pathway / activation of adenylate cyclase activity / negative regulation of blood pressure / positive regulation of gluconeogenesis / regulation of insulin secretion / cellular response to glucagon stimulus / adenylate cyclase activator activity / trans-Golgi network membrane / gluconeogenesis / response to activity / negative regulation of inflammatory response to antigenic stimulus / hormone activity / bone development / adenylate cyclase-modulating G protein-coupled receptor signaling pathway / G-protein beta/gamma-subunit complex binding / Olfactory Signaling Pathway / platelet aggregation / Activation of the phototransduction cascade / cognition / G beta:gamma signalling through PLC beta / Presynaptic function of Kainate receptors / Thromboxane signalling through TP receptor / G protein-coupled acetylcholine receptor signaling pathway / G-protein activation / Activation of G protein gated Potassium channels / Inhibition of voltage gated Ca2+ channels via Gbeta/gamma subunits / adenylate cyclase-activating G protein-coupled receptor signaling pathway / Prostacyclin signalling through prostacyclin receptor / G beta:gamma signalling through CDC42 / Glucagon signaling in metabolic regulation / G beta:gamma signalling through BTK / Synthesis, secretion, and inactivation of Glucagon-like Peptide-1 (GLP-1) / ADP signalling through P2Y purinoceptor 12 / Sensory perception of sweet, bitter, and umami (glutamate) taste / photoreceptor disc membrane / Glucagon-type ligand receptors / Adrenaline,noradrenaline inhibits insulin secretion / Vasopressin regulates renal water homeostasis via Aquaporins / G alpha (z) signalling events / Glucagon-like Peptide-1 (GLP1) regulates insulin secretion / transmembrane signaling receptor activity / ADORA2B mediated anti-inflammatory cytokines production / cellular response to catecholamine stimulus / ADP signalling through P2Y purinoceptor 1 / G beta:gamma signalling through PI3Kgamma / adenylate cyclase-activating dopamine receptor signaling pathway / Cooperation of PDCL (PhLP1) and TRiC/CCT in G-protein beta folding / sensory perception of smell / GPER1 signaling / Inactivation, recovery and regulation of the phototransduction cascade / cellular response to prostaglandin E stimulus / G-protein beta-subunit binding / heterotrimeric G-protein complex / glucose homeostasis / G alpha (12/13) signalling events / sensory perception of taste / extracellular vesicle / signaling receptor complex adaptor activity / Thrombin signalling through proteinase activated receptors (PARs) / positive regulation of cold-induced thermogenesis / G protein activity / GTPase binding / positive regulation of cytosolic calcium ion concentration / Ca2+ pathway / retina development in camera-type eye / High laminar flow shear stress activates signaling by PIEZO1 and PECAM1:CDH5:KDR in endothelial cells / fibroblast proliferation / phospholipase C-activating G protein-coupled receptor signaling pathway / G alpha (i) signalling events / Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement / G alpha (s) signalling events / secretory granule lumen / G alpha (q) signalling events / Ras protein signal transduction / positive regulation of ERK1 and ERK2 cascade / learning or memory / Extra-nuclear estrogen signaling / cell population proliferation
Similarity search - Function
Glucagon / GPCR, family 2, glucagon-like peptide-1 receptor / : / Glucagon/GIP/secretin/VIP / Peptide hormone / Glucagon / GIP / secretin / VIP family signature. / Glucagon like hormones / GPCR, family 2, glucagon-like peptide-1/glucagon receptor / G-protein coupled receptors family 2 signature 1. / : ...Glucagon / GPCR, family 2, glucagon-like peptide-1 receptor / : / Glucagon/GIP/secretin/VIP / Peptide hormone / Glucagon / GIP / secretin / VIP family signature. / Glucagon like hormones / GPCR, family 2, glucagon-like peptide-1/glucagon receptor / G-protein coupled receptors family 2 signature 1. / : / GPCR, family 2, extracellular hormone receptor domain / G-protein coupled receptors family 2 profile 1. / Domain present in hormone receptors / Hormone receptor domain / GPCR family 2, extracellular hormone receptor domain superfamily / G-protein coupled receptors family 2 signature 2. / GPCR, family 2, secretin-like, conserved site / GPCR, family 2, secretin-like / 7 transmembrane receptor (Secretin family) / GPCR, family 2-like / G-protein coupled receptors family 2 profile 2. / G-protein alpha subunit, group S / Guanine nucleotide binding protein (G-protein), alpha subunit / G protein alpha subunit, helical insertion / G-protein alpha subunit / G-alpha domain profile. / G protein alpha subunit / G-protein, gamma subunit / G-protein gamma subunit domain profile. / G-protein gamma-like domain / G-protein gamma-like domain superfamily / GGL domain / G protein gamma subunit-like motifs / GGL domain / Guanine nucleotide-binding protein, beta subunit / G-protein, beta subunit / G-protein beta WD-40 repeat / WD40 repeat, conserved site / Trp-Asp (WD) repeats signature. / Trp-Asp (WD) repeats profile. / Trp-Asp (WD) repeats circular profile. / WD domain, G-beta repeat / WD40 repeats / WD40 repeat / WD40-repeat-containing domain superfamily / WD40/YVTN repeat-like-containing domain superfamily / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Chem-QW7 / Pro-glucagon / Glucagon-like peptide 1 receptor / Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2 / Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1 / Guanine nucleotide-binding protein G(s) subunit alpha isoforms short
Similarity search - Component
Biological speciesHomo sapiens (human)
Lama glama (llama)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsSun, B. / Feng, D. / Bueno, A. / Kobilka, B. / Sloop, K.
CitationJournal: Nat Chem Biol / Year: 2020
Title: Structural insights into probe-dependent positive allosterism of the GLP-1 receptor.
Authors: Ana B Bueno / Bingfa Sun / Francis S Willard / Dan Feng / Joseph D Ho / David B Wainscott / Aaron D Showalter / Michal Vieth / Qi Chen / Cynthia Stutsman / Betty Chau / James Ficorilli / ...Authors: Ana B Bueno / Bingfa Sun / Francis S Willard / Dan Feng / Joseph D Ho / David B Wainscott / Aaron D Showalter / Michal Vieth / Qi Chen / Cynthia Stutsman / Betty Chau / James Ficorilli / Francisco J Agejas / Graham R Cumming / Alma Jiménez / Isabel Rojo / Tong Sun Kobilka / Brian K Kobilka / Kyle W Sloop /
Abstract: Drugs that promote the association of protein complexes are an emerging therapeutic strategy. We report discovery of a G protein-coupled receptor (GPCR) ligand that stabilizes an active state ...Drugs that promote the association of protein complexes are an emerging therapeutic strategy. We report discovery of a G protein-coupled receptor (GPCR) ligand that stabilizes an active state conformation by cooperatively binding both the receptor and orthosteric ligand, thereby acting as a 'molecular glue'. LSN3160440 is a positive allosteric modulator of the GLP-1R optimized to increase the affinity and efficacy of GLP-1(9-36), a proteolytic product of GLP-1(7-36). The compound enhances insulin secretion in a glucose-, ligand- and GLP-1R-dependent manner. Cryo-electron microscopy determined the structure of the GLP-1R bound to LSN3160440 in complex with GLP-1 and heterotrimeric G. The modulator binds high in the helical bundle at an interface between TM1 and TM2, allowing access to the peptide ligand. Pharmacological characterization showed strong probe dependence of LSN3160440 for GLP-1(9-36) versus oxyntomodulin that is driven by a single residue. Our findings expand protein-protein modulation drug discovery to uncompetitive, active state stabilizers for peptide hormone receptors.
History
DepositionDec 20, 2019Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jul 22, 2020Provider: repository / Type: Initial release
Revision 1.1Aug 5, 2020Group: Database references / Category: citation / citation_author
Item: _citation.pdbx_database_id_PubMed / _citation.title ..._citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID / _citation_author.name
Revision 1.2Sep 30, 2020Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first / _citation.page_last
Revision 1.3Oct 23, 2024Group: Data collection / Database references ...Data collection / Database references / Refinement description / Structure summary
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / em_3d_fitting_list / em_admin / pdbx_entry_details / pdbx_initial_refinement_model / pdbx_modification_feature
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id / _em_3d_fitting_list.source_name / _em_3d_fitting_list.type / _em_admin.last_update / _pdbx_entry_details.has_protein_modification

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Structure visualization

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Assembly

Deposited unit
R: Glucagon-like peptide 1 receptor
P: Glucagon-like peptide 1
A: Guanine nucleotide-binding protein G(s) subunit alpha isoforms short
B: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1
G: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2
N: Nanobody 35
hetero molecules


Theoretical massNumber of molelcules
Total (without water)163,1477
Polymers162,6676
Non-polymers4801
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area16170 Å2
ΔGint-78 kcal/mol
Surface area49280 Å2

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Components

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Glucagon-like peptide ... , 2 types, 2 molecules RP

#1: Protein Glucagon-like peptide 1 receptor / GLP-1R


Mass: 51233.109 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Homo sapiens (human) / References: UniProt: P43220
#2: Protein/peptide Glucagon-like peptide 1


Mass: 3359.699 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Homo sapiens (human) / References: UniProt: P01275

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Guanine nucleotide-binding protein ... , 3 types, 3 molecules ABG

#3: Protein Guanine nucleotide-binding protein G(s) subunit alpha isoforms short / Adenylate cyclase-stimulating G alpha protein


Mass: 44326.160 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: GNAS, GNAS1, GSP / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P63092
#4: Protein Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1 / Transducin beta chain 1


Mass: 38534.062 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: GNB1 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P62873
#5: Protein Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2 / G gamma-I


Mass: 7861.143 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: GNG2 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P59768

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Antibody / Non-polymers , 2 types, 2 molecules N

#6: Antibody Nanobody 35


Mass: 17352.498 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Lama glama (llama) / Production host: Escherichia coli (E. coli)
#7: Chemical ChemComp-QW7 / 1-[(1R)-1-(2,6-dichloro-3-methoxyphenyl)ethyl]-6-{2-[(2R)-piperidin-2-yl]phenyl}-1H-benzimidazole


Mass: 480.429 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C27H27Cl2N3O / Feature type: SUBJECT OF INVESTIGATION

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Details

Has ligand of interestY
Has protein modificationY

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

Component
IDNameTypeEntity IDParent-IDSource
1Cryo-EM structure of the Glucagon-like peptide-1 receptor in complex with G protein, GLP-1 peptide and a positive allosteric modulatorCOMPLEX#1-#60RECOMBINANT
3Guanine nucleotide-binding protein G(s) subunit alpha isoforms short, Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1, Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2COMPLEX#3-#51RECOMBINANT
2Glucagon-like peptide 1 receptor, Glucagon-like peptide 1COMPLEX#1-#21RECOMBINANT
4Nanobody 35COMPLEX#61RECOMBINANT
Molecular weightValue: 0.15 MDa / Experimental value: NO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
12Homo sapiens (human)9606
23Homo sapiens (human)9606
34Lama glama (llama)9844
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
23Spodoptera frugiperda (fall armyworm)7108
12synthetic construct (others)32630
34Escherichia coli (E. coli)562
Buffer solutionpH: 7.5
SpecimenConc.: 10 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 200 divisions/in. / Grid type: Quantifoil, UltrAuFoil, R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 293 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Cs: 2.7 mm
Image recordingAverage exposure time: 10.8 sec. / Electron dose: 81.56 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k)
EM imaging opticsEnergyfilter name: GIF Quantum LS / Energyfilter slit width: 20 eV
Image scansMovie frames/image: 54

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Processing

Software
NameVersionClassification
phenix.real_space_refine1.13_2998refinement
PHENIX1.13_2998refinement
EM software
IDNameVersionCategory
4RELION3CTF correction
7PHENIX1.13model fitting
12RELION33D reconstruction
13PHENIX1.13model refinement
CTF correctionType: NONE
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 323427 / Symmetry type: POINT
Atomic model buildingB value: 72.5 / Protocol: FLEXIBLE FIT
Atomic model buildingPDB-ID: 5VAI

5vai


Accession code: 5VAI / Source name: PDB / Type: experimental model
RefinementStereochemistry target values: GeoStd + Monomer Library
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00639283
ELECTRON MICROSCOPYf_angle_d0.990112605
ELECTRON MICROSCOPYf_chiral_restr0.05981404
ELECTRON MICROSCOPYf_plane_restr0.00671599
ELECTRON MICROSCOPYf_dihedral_angle_d12.74625457

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