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- PDB-8cqb: Cryo-EM structure of the human GBP1 dimer bound to GDP-AlF3 -

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Basic information

Entry
Database: PDB / ID: 8cqb
TitleCryo-EM structure of the human GBP1 dimer bound to GDP-AlF3
ComponentsGuanylate-binding protein 1
KeywordsIMMUNE SYSTEM / Cell-autonomous immunity / intracellular pathogens / GTPase
Function / homology
Function and homology information


GDP phosphatase activity / non-canonical inflammasome complex assembly / protein localization to vacuole / negative regulation of substrate adhesion-dependent cell spreading / symbiont cell surface / cytolysis in another organism / positive regulation of pyroptosis / vesicle membrane / negative regulation of protein localization to plasma membrane / negative regulation of interleukin-2 production ...GDP phosphatase activity / non-canonical inflammasome complex assembly / protein localization to vacuole / negative regulation of substrate adhesion-dependent cell spreading / symbiont cell surface / cytolysis in another organism / positive regulation of pyroptosis / vesicle membrane / negative regulation of protein localization to plasma membrane / negative regulation of interleukin-2 production / negative regulation of T cell receptor signaling pathway / spectrin binding / cytokine binding / defense response to protozoan / Hydrolases; Acting on acid anhydrides; In phosphorus-containing anhydrides / cellular response to interleukin-1 / regulation of protein localization to plasma membrane / regulation of calcium-mediated signaling / G protein activity / lipopolysaccharide binding / Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement / Hsp90 protein binding / cytoplasmic vesicle membrane / negative regulation of ERK1 and ERK2 cascade / cellular response to type II interferon / GDP binding / Interferon gamma signaling / actin cytoskeleton / cellular response to tumor necrosis factor / actin binding / cytoplasmic vesicle / defense response to virus / defense response to bacterium / Golgi membrane / innate immune response / GTPase activity / GTP binding / Golgi apparatus / enzyme binding / protein homodimerization activity / extracellular region / identical protein binding / plasma membrane / cytosol / cytoplasm
Similarity search - Function
Guanylate-binding protein, C-terminal / Guanylate-binding protein/Atlastin, C-terminal / Guanylate-binding protein, C-terminal domain / Guanylate-binding protein, N-terminal / Guanylate-binding protein, C-terminal domain superfamily / Guanylate-binding protein, N-terminal domain / GB1/RHD3-type guanine nucleotide-binding (G) domain / GB1/RHD3-type guanine nucleotide-binding (G) domain profile. / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
ALUMINUM FLUORIDE / GUANOSINE-5'-DIPHOSPHATE / Guanylate-binding protein 1
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.7 Å
AuthorsKuhm, T.I. / Jakobi, A.J.
Funding supportEuropean Union, Netherlands, 2items
OrganizationGrant numberCountry
European Research Council (ERC)852880European Union
Netherlands Organisation for Scientific Research (NWO)NWO.STU.018-2.007 Netherlands
CitationJournal: To Be Published
Title: Cryo-EM structure of the human GBP1 dimer bound to GDP-AlF3
Authors: Kuhm, T.I. / de Agrela Pinto, C. / Taisne, C. / Huber, S. / Giannopolou, N. / Pardon, E. / Steyaert, J. / Jakobi, A.J.
History
DepositionMar 4, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Mar 13, 2024Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Guanylate-binding protein 1
B: Guanylate-binding protein 1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)137,1468
Polymers136,0432
Non-polymers1,1036
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: light scattering, native gel electrophoresis, electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area9830 Å2
ΔGint-59 kcal/mol
Surface area42530 Å2

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Components

#1: Protein Guanylate-binding protein 1 / GTP-binding protein 1 / GBP-1 / HuGBP-1 / Guanine nucleotide-binding protein 1 / Interferon-induced ...GTP-binding protein 1 / GBP-1 / HuGBP-1 / Guanine nucleotide-binding protein 1 / Interferon-induced guanylate-binding protein 1


Mass: 68021.617 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: GBP1 / Production host: Escherichia coli (E. coli)
References: UniProt: P32455, Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement
#2: Chemical ChemComp-GDP / GUANOSINE-5'-DIPHOSPHATE / Guanosine diphosphate


Type: RNA linking / Mass: 443.201 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C10H15N5O11P2 / Feature type: SUBJECT OF INVESTIGATION / Comment: GDP, energy-carrying molecule*YM
#3: Chemical ChemComp-AF3 / ALUMINUM FLUORIDE / Aluminium fluoride


Mass: 83.977 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: AlF3 / Feature type: SUBJECT OF INVESTIGATION
#4: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Mg / Feature type: SUBJECT OF INVESTIGATION
Has ligand of interestY

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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: Homodimeric complex of GBP1 stabilised by GDP-AlF3 / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Molecular weightValue: 0.134 MDa / Experimental value: YES
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Escherichia coli (E. coli) / Strain: BL21(DE3)pLysS / Plasmid: pETM14
Buffer solutionpH: 7.4
Details: 5u mM HEPES pH7.4, 150 mM NaCl, 1 mM DTT, 200 uM GDP, 10 mM NaF, 300 uM AlCl3, 5 mM MgCl2
Buffer component
IDConc.NameFormulaBuffer-ID
150 mMHEPESC8H18N2O4S1
2150 mMSodium chlorideNaClSodium chloride1
31 mMDithiothreitolC4H10O2S21
4200 uMGuanosine diphosphateC10H15N5O11P21
510 mMSodium fluorideNaF1
6300 uMAluminium chlorideAlCl31
75 mMMagnesium chlorideMgCl21
SpecimenConc.: 0.7 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil
VitrificationInstrument: LEICA EM GP / Cryogen name: ETHANE / Humidity: 99 % / Chamber temperature: 22 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 FIELDBright-field microscopy / Nominal magnification: 105000 X / Nominal defocus max: 2200 nm / Nominal defocus min: 600 nm / Cs: 2.7 mm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 60 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 2 / Num. of real images: 5214
EM imaging opticsEnergyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV
Image scansWidth: 5760 / Height: 4092

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Processing

SoftwareName: UCSF ChimeraX / Version: 1.3/v9 / Classification: model building / URL: https://www.rbvi.ucsf.edu/chimerax/ / Os: macOS / Type: package
EM software
IDNameVersionCategory
1Topazparticle selection
2EPU2.8.1image acquisition
4cryoSPARC3.1CTF correction
7Coot0.9.5model fitting
11cryoSPARC3.3classification
12cryoSPARC3.33D reconstruction
19REFMAC5model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 447651
Details: 447,651 particles were used to perform ab initio reconstruction to generate five different models. Three classes were selected for heterogeneous refinements without imposing symmetry. A ...Details: 447,651 particles were used to perform ab initio reconstruction to generate five different models. Three classes were selected for heterogeneous refinements without imposing symmetry. A single class with 147095 particles was used for non-uniform refinement.
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 147095 / Symmetry type: POINT
Atomic model buildingProtocol: OTHER / Space: RECIPROCAL / Target criteria: Real-space cross correlation
Atomic model building

3D fitting-ID: 1 / Source name: PDB / Type: experimental model

IDPDB-IDPdb chain-IDAccession codeChain-IDChain residue rangeInitial refinement model-IDPdb chain residue range
11FN5

1fn5

A1FN5A323-4781323-478
22B92

2b92

A2B92A6-30426-304
32B92

2b92

B2B92B6-30426-304

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