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- PDB-8flr: Human PTH1R in complex with PTHrP and Gs -

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Basic information

Entry
Database: PDB / ID: 8flr
TitleHuman PTH1R in complex with PTHrP and Gs
Components
  • (Guanine nucleotide-binding protein ...) x 3
  • Nanobody35
  • PTHrP[1-36]
  • Parathyroid hormone/parathyroid hormone-related peptide receptor
KeywordsMEMBRANE PROTEIN / GPCR / agonist / hormone
Function / homology
Function and homology information


negative regulation of chondrocyte development / regulation of chondrocyte differentiation / parathyroid hormone receptor activity / cAMP metabolic process / G protein-coupled peptide receptor activity / negative regulation of chondrocyte differentiation / Class B/2 (Secretin family receptors) / osteoblast development / positive regulation of inositol phosphate biosynthetic process / peptide hormone receptor binding ...negative regulation of chondrocyte development / regulation of chondrocyte differentiation / parathyroid hormone receptor activity / cAMP metabolic process / G protein-coupled peptide receptor activity / negative regulation of chondrocyte differentiation / Class B/2 (Secretin family receptors) / osteoblast development / positive regulation of inositol phosphate biosynthetic process / peptide hormone receptor binding / bone mineralization / peptide hormone binding / PKA activation in glucagon signalling / G protein-coupled receptor signaling pathway, coupled to cyclic nucleotide second messenger / hair follicle placode formation / developmental growth / D1 dopamine receptor binding / intracellular transport / epidermis development / positive regulation of cAMP-mediated signaling / chondrocyte differentiation / Hedgehog 'off' state / adenylate cyclase-activating adrenergic receptor signaling pathway / cell maturation / bone resorption / activation of adenylate cyclase activity / adenylate cyclase activator activity / trans-Golgi network membrane / skeletal system development / female pregnancy / G-protein beta/gamma-subunit complex binding / Olfactory Signaling Pathway / Activation of the phototransduction cascade / bone development / G beta:gamma signalling through PLC beta / Presynaptic function of Kainate receptors / adenylate cyclase-modulating G protein-coupled receptor signaling pathway / Thromboxane signalling through TP receptor / G-protein activation / G protein-coupled acetylcholine receptor signaling pathway / Activation of G protein gated Potassium channels / Inhibition of voltage gated Ca2+ channels via Gbeta/gamma subunits / Prostacyclin signalling through prostacyclin receptor / Glucagon signaling in metabolic regulation / G beta:gamma signalling through CDC42 / adenylate cyclase-activating G protein-coupled receptor signaling pathway / hormone activity / ADP signalling through P2Y purinoceptor 12 / G beta:gamma signalling through BTK / Synthesis, secretion, and inactivation of Glucagon-like Peptide-1 (GLP-1) / Sensory perception of sweet, bitter, and umami (glutamate) taste / cognition / photoreceptor disc membrane / platelet aggregation / Adrenaline,noradrenaline inhibits insulin secretion / positive regulation of GTPase activity / Glucagon-type ligand receptors / Vasopressin regulates renal water homeostasis via Aquaporins / G alpha (z) signalling events / cellular response to catecholamine stimulus / Glucagon-like Peptide-1 (GLP1) regulates insulin secretion / intracellular calcium ion homeostasis / ADORA2B mediated anti-inflammatory cytokines production / sensory perception of taste / ADP signalling through P2Y purinoceptor 1 / adenylate cyclase-activating dopamine receptor signaling pathway / G beta:gamma signalling through PI3Kgamma / cellular response to prostaglandin E stimulus / Cooperation of PDCL (PhLP1) and TRiC/CCT in G-protein beta folding / GPER1 signaling / G-protein beta-subunit binding / Inactivation, recovery and regulation of the phototransduction cascade / heterotrimeric G-protein complex / : / G alpha (12/13) signalling events / extracellular vesicle / sensory perception of smell / signaling receptor complex adaptor activity / Thrombin signalling through proteinase activated receptors (PARs) / retina development in camera-type eye / GTPase binding / cell-cell signaling / phospholipase C-activating G protein-coupled receptor signaling pathway / Ca2+ pathway / positive regulation of cold-induced thermogenesis / G alpha (i) signalling events / fibroblast proliferation / G alpha (s) signalling events / G alpha (q) signalling events / regulation of gene expression / basolateral plasma membrane / in utero embryonic development / cell population proliferation / Ras protein signal transduction / Extra-nuclear estrogen signaling / cell surface receptor signaling pathway / receptor complex / apical plasma membrane / G protein-coupled receptor signaling pathway / lysosomal membrane
Similarity search - Function
Parathyroid hormone-related protein / Parathyroid hormone/parathyroid hormone-related protein / Parathyroid hormone family / Parathyroid hormone family signature. / Parathyroid hormone / GPCR, family 2, parathyroid hormone receptor / G-protein coupled receptors family 2 signature 1. / Hormone receptor domain / GPCR, family 2, extracellular hormone receptor domain / G-protein coupled receptors family 2 profile 1. ...Parathyroid hormone-related protein / Parathyroid hormone/parathyroid hormone-related protein / Parathyroid hormone family / Parathyroid hormone family signature. / Parathyroid hormone / GPCR, family 2, parathyroid hormone receptor / G-protein coupled receptors family 2 signature 1. / Hormone receptor domain / GPCR, family 2, extracellular hormone receptor domain / G-protein coupled receptors family 2 profile 1. / Domain present in hormone receptors / 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 / G-alpha domain profile. / Guanine nucleotide binding protein (G-protein), alpha subunit / G protein alpha subunit, helical insertion / G-protein alpha subunit / G protein alpha subunit / G-protein, gamma subunit / G-protein gamma subunit domain profile. / GGL domain / G-protein gamma-like domain superfamily / G-protein gamma-like domain / GGL domain / G protein gamma subunit-like motifs / 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
Parathyroid hormone-related protein / 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 / Parathyroid hormone/parathyroid hormone-related peptide receptor
Similarity search - Component
Biological speciesHomo sapiens (human)
Lama glama (llama)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.94 Å
AuthorsCary, B.P. / Belousoff, M.J. / Piper, S.J. / Wootten, D. / Sexton, P.M.
Funding support Australia, 4items
OrganizationGrant numberCountry
Australian Research Council (ARC)IC200100052 Australia
National Health and Medical Research Council (NHMRC, Australia)1155302 Australia
National Health and Medical Research Council (NHMRC, Australia)1154434 Australia
National Health and Medical Research Council (NHMRC, Australia)1150083 Australia
CitationJournal: Structure / Year: 2023
Title: Molecular insights into peptide agonist engagement with the PTH receptor.
Authors: Brian P Cary / Elliot J Gerrard / Matthew J Belousoff / Madeleine M Fletcher / Yan Jiang / Isabella C Russell / Sarah J Piper / Denise Wootten / Patrick M Sexton /
Abstract: The parathyroid hormone (PTH) 1 receptor (PTH1R) is a G protein-coupled receptor (GPCR) that regulates skeletal development and calcium homeostasis. Here, we describe cryo-EM structures of the PTH1R ...The parathyroid hormone (PTH) 1 receptor (PTH1R) is a G protein-coupled receptor (GPCR) that regulates skeletal development and calcium homeostasis. Here, we describe cryo-EM structures of the PTH1R in complex with fragments of the two hormones, PTH and PTH-related protein, the drug abaloparatide, as well as the engineered tool compounds, long-acting PTH (LA-PTH) and the truncated peptide, M-PTH(1-14). We found that the critical N terminus of each agonist engages the transmembrane bundle in a topologically similar fashion, reflecting similarities in measures of Gαs activation. The full-length peptides induce subtly different extracellular domain (ECD) orientations relative to the transmembrane domain. In the structure bound to M-PTH, the ECD is unresolved, demonstrating that the ECD is highly dynamic when unconstrained by a peptide. High resolutions enabled identification of water molecules near peptide and G protein binding sites. Our results illuminate the action of orthosteric agonists of the PTH1R.
History
DepositionDec 22, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Apr 26, 2023Provider: repository / Type: Initial release
Revision 1.1May 17, 2023Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _citation_author.identifier_ORCID
Revision 1.2Jun 14, 2023Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first / _citation.page_last

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
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: Nanobody35
P: PTHrP[1-36]
R: Parathyroid hormone/parathyroid hormone-related peptide receptor


Theoretical massNumber of molelcules
Total (without water)177,1426
Polymers177,1426
Non-polymers00
Water18010
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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

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


Mass: 45683.434 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: Trichoplusia ni (cabbage looper) / References: UniProt: P63092
#2: Protein Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1 / Transducin beta chain 1


Mass: 37413.863 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: GNB1 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P62873
#3: Protein Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2 / G gamma-I


Mass: 6375.332 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: GNG2 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P59768

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Antibody / Protein/peptide / Protein / Non-polymers , 4 types, 13 molecules NPR

#4: Antibody Nanobody35


Mass: 13885.439 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)
#5: Protein/peptide PTHrP[1-36]


Mass: 4269.908 Da / Num. of mol.: 1 / Fragment: UNP residues 37-72 / Source method: obtained synthetically / Source: (synth.) Homo sapiens (human) / References: UniProt: P12272
#6: Protein Parathyroid hormone/parathyroid hormone-related peptide receptor / PTH/PTHrP type I receptor / PTH/PTHr receptor / Parathyroid hormone 1 receptor / PTH1 receptor


Mass: 69513.758 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: PTH1R, PTHR, PTHR1 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q03431
#7: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 10 / Source method: isolated from a natural source / Formula: H2O

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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

ComponentName: PTHrP bound to the PTH1R in complex with G protein / Type: COMPLEX / Entity ID: #1-#6 / Source: MULTIPLE SOURCES
Molecular weightExperimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Trichoplusia ni (cabbage looper)
Buffer solutionpH: 7.4 / Details: Buffer also contained 2 micromolar PTHrP peptide.
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMHEPESC8H18N2O4S1
2100 mMSodium ChlorideNaClSodium chloride1
32 mMMagnesium ChlorideMgCl21
40.1 mMTCEPC9H15O6P1
50.01 percentLMNGC47H88O221
60.00006 percentCHSC31H50O41
SpecimenConc.: 3.66 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: 20 mA current, negative polarity / Grid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K

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Electron microscopy imaging

MicroscopyModel: TFS GLACIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 1300 nm / Nominal defocus min: 600 nm / Cs: 2.7 mm
Specimen holderCryogen: NITROGEN
Image recordingAverage exposure time: 7.39 sec. / Electron dose: 50 e/Å2 / Film or detector model: FEI FALCON IV (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 4553

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Processing

SoftwareName: PHENIX / Version: 1.19.2_4158: / Classification: refinement
EM software
IDNameVersionCategory
4CTFFIND4.1.14CTF correction
9cryoSPARC3.1.2initial Euler assignment
10cryoSPARC3.3.2final Euler assignment
12cryoSPARC3.3.23D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 2140434
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 2.94 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 395975 / Symmetry type: POINT
Atomic model building
IDPDB-IDPdb chain-ID 3D fitting-ID
16NBF

6nbf

R1
26X18

6x18

1
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0039146
ELECTRON MICROSCOPYf_angle_d0.50212412
ELECTRON MICROSCOPYf_dihedral_angle_d4.2121247
ELECTRON MICROSCOPYf_chiral_restr0.041389
ELECTRON MICROSCOPYf_plane_restr0.0041580

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