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- PDB-7joa: 2:1 cGAS-nucleosome complex -

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Basic information

Entry
Database: PDB / ID: 7joa
Title2:1 cGAS-nucleosome complex
Components
  • (DNA (145-MER)) x 2
  • Cyclic GMP-AMP synthase
  • Histone H2A type 1
  • Histone H2B type 1-C/E/F/G/I
  • Histone H3.2
  • Histone H4
KeywordsDna Binding Protein/DNA/Transferase / cGAS / nucleosome / cyclic GMP-AMP synthase / Dna Binding Protein-DNA-Transferase complex
Function / homology
Function and homology information


cyclic GMP-AMP synthase / positive regulation of cGMP-mediated signaling / cyclic-GMP-AMP synthase activity / regulation of immunoglobulin production / paracrine signaling / regulation of type I interferon production / negative regulation of double-strand break repair via homologous recombination / regulation of T cell activation / positive regulation of cAMP-mediated signaling / cellular response to exogenous dsRNA ...cyclic GMP-AMP synthase / positive regulation of cGMP-mediated signaling / cyclic-GMP-AMP synthase activity / regulation of immunoglobulin production / paracrine signaling / regulation of type I interferon production / negative regulation of double-strand break repair via homologous recombination / regulation of T cell activation / positive regulation of cAMP-mediated signaling / cellular response to exogenous dsRNA / negative regulation of megakaryocyte differentiation / positive regulation of defense response to virus by host / CENP-A containing nucleosome assembly / activation of innate immune response / DNA replication-independent nucleosome assembly / telomere capping / interleukin-7-mediated signaling pathway / chromatin silencing / determination of adult lifespan / positive regulation of type I interferon production / telomere organization / DNA replication-dependent nucleosome assembly / innate immune response in mucosa / nuclear nucleosome / rDNA heterochromatin assembly / negative regulation of gene expression, epigenetic / regulation of gene silencing by miRNA / nuclear chromosome / DNA-templated transcription, initiation / regulation of megakaryocyte differentiation / phosphatidylinositol-4,5-bisphosphate binding / nucleosome assembly / nucleosome / site of double-strand break / positive regulation of cellular senescence / double-strand break repair via nonhomologous end joining / defense response to virus / double-stranded DNA binding / chromatin organization / regulation of immune response / nuclear chromosome, telomeric region / antibacterial humoral response / antimicrobial humoral immune response mediated by antimicrobial peptide / blood coagulation / protein ubiquitination / protein heterodimerization activity / defense response to Gram-positive bacterium / nuclear chromatin / DNA repair / protein domain specific binding / GTP binding / cellular response to DNA damage stimulus / chromatin binding / cellular protein metabolic process / innate immune response / host cell nucleus / enzyme binding / protein-containing complex / RNA binding / DNA binding / extracellular space / extracellular exosome / membrane / extracellular region / nucleoplasm / ATP binding / identical protein binding / plasma membrane / metal ion binding / nucleus / cytosol
Histone H2B / TATA box binding protein associated factor (TAF) / CENP-T/Histone H4, histone fold / Histone H2A conserved site / Histone H2A, C-terminal domain / Mab-21 domain / Histone H4, conserved site / Histone H3/CENP-A / Histone-fold / Histone H4 ...Histone H2B / TATA box binding protein associated factor (TAF) / CENP-T/Histone H4, histone fold / Histone H2A conserved site / Histone H2A, C-terminal domain / Mab-21 domain / Histone H4, conserved site / Histone H3/CENP-A / Histone-fold / Histone H4 / Histone H2A / Histone H2A/H2B/H3
Histone H2A type 1 / Histone H4 / Histone H2B type 1-C/E/F/G/I / Histone H3.2 / Cyclic GMP-AMP synthase
Biological speciesHomo sapiens (human)
synthetic construct (others)
Mus musculus (house mouse)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsBoyer, J.A. / Spangler, C.J. / Strauss, J.D. / Cesmat, A.P. / Liu, P. / McGinty, R.K. / Zhang, Q.
Funding support United States, 2items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)1R35GM133498 United States
National Institutes of Health/National Cancer Institute (NIH/NCI)R21CA234979 United States
CitationJournal: Science / Year: 2020
Title: Structural basis of nucleosome-dependent cGAS inhibition.
Authors: Joshua A Boyer / Cathy J Spangler / Joshua D Strauss / Andrew P Cesmat / Pengda Liu / Robert K McGinty / Qi Zhang /
Abstract: Cyclic GMP-AMP synthase (cGAS) recognizes cytosolic foreign or damaged DNA to activate the innate immune response to infection, inflammatory diseases, and cancer. In contrast, cGAS reactivity against ...Cyclic GMP-AMP synthase (cGAS) recognizes cytosolic foreign or damaged DNA to activate the innate immune response to infection, inflammatory diseases, and cancer. In contrast, cGAS reactivity against self-DNA in the nucleus is suppressed by chromatin tethering. We report a 3.3-angstrom-resolution cryo-electron microscopy structure of cGAS in complex with the nucleosome core particle. The structure reveals that cGAS employs two conserved arginines to anchor to the nucleosome acidic patch. The nucleosome binding interface exclusively occupies the strong dsDNA binding surface on cGAS and sterically prevents cGAS from oligomerizing into the functionally active 2:2 cGAS-dsDNA state. These findings provide a structural basis for how cGAS maintains an inhibited state in the nucleus and further exemplify the role of the nucleosome in regulating diverse nuclear protein functions.
Validation Report
SummaryFull reportAbout validation report
History
DepositionAug 6, 2020Deposition site: RCSB / Processing site: RCSB
Revision 1.0Sep 16, 2020Provider: repository / Type: Initial release
Revision 1.1Sep 23, 2020Group: Database references / Category: citation / citation_author
Item: _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID

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Structure visualization

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  • Deposited structure unit
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Structure viewerMolecule:
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Assembly

Deposited unit
A: Histone H3.2
B: Histone H4
C: Histone H2A type 1
D: Histone H2B type 1-C/E/F/G/I
E: Histone H3.2
F: Histone H4
G: Histone H2A type 1
H: Histone H2B type 1-C/E/F/G/I
I: DNA (145-MER)
J: DNA (145-MER)
K: Cyclic GMP-AMP synthase
hetero molecules


Theoretical massNumber of molelcules
Total (without water)243,02312
Polymers242,95711
Non-polymers651
Water0
1


TypeNameSymmetry operationNumber
identity operation1_5551
Noncrystallographic symmetry (NCS)NCS domain:
IDEns-IDDetails
d_1ens_1chain "G"
d_2ens_1chain "C"
d_1ens_2chain "D"
d_2ens_2chain "H"

NCS domain segments:
Dom-IDComponent-IDEns-IDBeg label comp-IDEnd label comp-IDLabel asym-IDLabel seq-ID
d_11ens_1ALALYSG1 - 109
d_21ens_1ALALYSC1 - 109
d_11ens_2ARGSERD1 - 94
d_21ens_2ARGSERH1 - 94

NCS ensembles:
ID
ens_1
ens_2

NCS oper:
IDCodeMatrixVector
1given(-0.999946616659, 0.0091768519037, -0.00474860210389), (-0.00916331043712, -0.99995390993, -0.00286561658816), (-0.00477468057953, -0.00282195069675, 0.999984619392)172.989553801, 91.9551941256, 0.626130540196
2given(-0.999991465495, -0.000659074979583, -0.00407854850795), (0.000656337664566, -0.999999558515, 0.000672451377636), (-0.00407898990321, 0.000669768733594, 0.999991456589)173.208597822, 90.8717263617, 0.298977679173

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Components

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Protein , 5 types, 9 molecules AEBFCGDHK

#1: Protein Histone H3.2 / H3-clustered histone 13 / H3-clustered histone 14 / H3-clustered histone 15 / Histone H3/m / Histone H3/o


Mass: 15421.101 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human)
Gene: H3C15, HIST2H3A, H3C14, H3F2, H3FM, HIST2H3C, H3C13, HIST2H3D
Production host: Escherichia coli BL21(DE3) (bacteria) / Variant (production host): pLysS / References: UniProt: Q71DI3
#2: Protein Histone H4 /


Mass: 11394.426 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human)
Gene: HIST1H4A, H4/A, H4FA, HIST1H4B, H4/I, H4FI, HIST1H4C, H4/G, H4FG, HIST1H4D, H4/B, H4FB, HIST1H4E, H4/J, H4FJ, HIST1H4F, H4/C, H4FC, HIST1H4H, H4/H, H4FH, HIST1H4I, H4/M, H4FM, HIST1H4J, H4/E, ...Gene: HIST1H4A, H4/A, H4FA, HIST1H4B, H4/I, H4FI, HIST1H4C, H4/G, H4FG, HIST1H4D, H4/B, H4FB, HIST1H4E, H4/J, H4FJ, HIST1H4F, H4/C, H4FC, HIST1H4H, H4/H, H4FH, HIST1H4I, H4/M, H4FM, HIST1H4J, H4/E, H4FE, HIST1H4K, H4/D, H4FD, HIST1H4L, H4/K, H4FK, HIST2H4A, H4/N, H4F2, H4FN, HIST2H4, HIST2H4B, H4/O, H4FO, HIST4H4
Production host: Escherichia coli BL21(DE3) (bacteria) / Variant (production host): pLysS / References: UniProt: P62805
#3: Protein Histone H2A type 1 / H2A.1 / Histone H2A/ptl


Mass: 13990.342 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human)
Gene: H2AC11, H2AFP, HIST1H2AG, H2AC13, H2AFC, HIST1H2AI, H2AC15, H2AFD, HIST1H2AK, H2AC16, H2AFI, HIST1H2AL, H2AC17, H2AFN, HIST1H2AM
Production host: Escherichia coli BL21(DE3) (bacteria) / Variant (production host): pLysS / References: UniProt: P0C0S8
#4: Protein Histone H2B type 1-C/E/F/G/I / Histone H2B.1 A / Histone H2B.a / H2B/a / Histone H2B.g / H2B/g / Histone H2B.h / H2B/h / Histone ...Histone H2B.1 A / Histone H2B.a / H2B/a / Histone H2B.g / H2B/g / Histone H2B.h / H2B/h / Histone H2B.k / H2B/k / Histone H2B.l / H2B/l


Mass: 13806.018 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human)
Gene: H2BC4, H2BFL, HIST1H2BC, H2BC6, H2BFH, HIST1H2BE, H2BC7, H2BFG, HIST1H2BF, H2BC8, H2BFA, HIST1H2BG, H2BC10, H2BFK, HIST1H2BI
Production host: Escherichia coli BL21(DE3) (bacteria) / Variant (production host): pLysS / References: UniProt: P62807
#7: Protein Cyclic GMP-AMP synthase / / m-cGAS / 2'3'-cGAMP synthase / Mab-21 domain-containing protein 1


Mass: 42984.664 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mus musculus (house mouse) / Gene: Cgas, Mb21d1 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q8C6L5, cyclic GMP-AMP synthase

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DNA chain , 2 types, 2 molecules IJ

#5: DNA chain DNA (145-MER)


Mass: 45610.043 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) synthetic construct (others) / Production host: Escherichia coli HB101 (bacteria)
#6: DNA chain DNA (145-MER)


Mass: 45138.770 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) synthetic construct (others) / Production host: Escherichia coli HB101 (bacteria)

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Non-polymers , 1 types, 1 molecules

#8: Chemical ChemComp-ZN / ZINC ION / Zinc


Mass: 65.409 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Zn

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Details

Has ligand of interestN

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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: 2:1 cGAS-nucleosome complex / Type: COMPLEX / Details: cGAS bound to the nucleosome in a 2:1 ratio / Entity ID: #1-#7 / Source: MULTIPLE SOURCES
Molecular weightValue: 0.29 MDa / Experimental value: NO
Source (natural)Organism: Homo sapiens (human)
Buffer solutionpH: 7.5
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMHEPESC8H18N2O4S1
2150 mMsodium chlorideNaClSodium chloride1
31 mMTCEPC9H15O6P1
SpecimenConc.: 1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: Instrument: Pelco easiGlow / Grid material: COPPER / Grid mesh size: 400 divisions/in. / Grid type: Quantifoil
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 95 % / Chamber temperature: 293 K

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Electron microscopy imaging

Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company
MicroscopyModel: FEI TALOS ARCTICA
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy
Image recordingElectron dose: 53 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 2100

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Processing

Software
NameVersionClassification
phenix.real_space_refine1.18_3861refinement
PHENIX1.18_3861refinement
EM software
IDNameVersionCategory
2SerialEMimage acquisition
4CTFFIND4.1CTF correction
7PHENIX1.18model fitting
9PHENIX1.18model refinement
10RELION3.1-betainitial Euler assignment
11RELION3.1-betafinal Euler assignment
12RELION3.1-betaclassification
13RELION3.1-beta3D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 433445
SymmetryPoint symmetry: C2 (2 fold cyclic)
3D reconstructionResolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 45587 / Symmetry type: POINT
Atomic model buildingSpace: REAL
Atomic model building
IDPDB-ID3D fitting-ID
16FQ51
24K8V1
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 51.24 Å2
Refine LS restraints
Refinement-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.008615861
ELECTRON MICROSCOPYf_angle_d0.771822634
ELECTRON MICROSCOPYf_chiral_restr0.04462547
ELECTRON MICROSCOPYf_plane_restr0.00571861
ELECTRON MICROSCOPYf_dihedral_angle_d27.03546473
Refine LS restraints NCS
Ens-IDDom-IDAuth asym-IDRefinement-IDTypeRms dev position (Å)
ens_1d_2GELECTRON MICROSCOPYNCS constraints0.00120437799177
ens_2d_2DELECTRON MICROSCOPYNCS constraints0.000909689883197

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