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- PDB-8ecc: Structure of Ternary Complex of cGAS with dsDNA and Bound 5-pppI(... -

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Basic information

Entry
Database: PDB / ID: 8ecc
TitleStructure of Ternary Complex of cGAS with dsDNA and Bound 5-pppI(2,5)pA
Components
  • Cyclic GMP-AMP synthase
  • Palindromic DNA18
KeywordsTRANSFERASE/DNA / Transferase-DNA complex / cGAS / 5-pppI(2 / 5)pA / IMMUNE SYSTEM / TRANSFERASE
Function / homology
Function and homology information


2',3'-cyclic GMP-AMP synthase activity / cyclic GMP-AMP synthase / regulation of type I interferon production / paracrine signaling / poly-ADP-D-ribose modification-dependent protein binding / regulation of immunoglobulin production / negative regulation of DNA repair / cGAS/STING signaling pathway / regulation of T cell activation / : ...2',3'-cyclic GMP-AMP synthase activity / cyclic GMP-AMP synthase / regulation of type I interferon production / paracrine signaling / poly-ADP-D-ribose modification-dependent protein binding / regulation of immunoglobulin production / negative regulation of DNA repair / cGAS/STING signaling pathway / regulation of T cell activation / : / negative regulation of cGAS/STING signaling pathway / cellular response to exogenous dsRNA / positive regulation of type I interferon production / regulation of immune response / negative regulation of double-strand break repair via homologous recombination / : / nucleosome binding / positive regulation of defense response to virus by host / phosphatidylinositol-4,5-bisphosphate binding / activation of innate immune response / determination of adult lifespan / molecular condensate scaffold activity / positive regulation of cellular senescence / site of double-strand break / double-stranded DNA binding / defense response to virus / nuclear body / innate immune response / DNA repair / DNA damage response / chromatin binding / GTP binding / protein homodimerization activity / DNA binding / ATP binding / metal ion binding / nucleus / plasma membrane / cytosol
Similarity search - Function
Mab-21-like, nucleotidyltransferase domain / Mab-21 protein nucleotidyltransferase domain / Mab-21-like / Mab-21-like, HhH/H2TH-like domain / Mab-21 protein HhH/H2TH-like domain / Mab-21
Similarity search - Domain/homology
ADENOSINE-5'-TRIPHOSPHATE / Chem-VWX / DNA / DNA (> 10) / Cyclic GMP-AMP synthase
Similarity search - Component
Biological speciesMus musculus (house mouse)
synthetic construct (others)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.44 Å
AuthorsWu, S. / Sohn, J.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS) United States
CitationJournal: To Be Published
Title: Structure of Ternary Complex of cGAS with dsDNA and Bound 5-pppI(2,5)pA
Authors: Wu, S. / Sohn, J.
History
DepositionSep 1, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jan 17, 2024Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Cyclic GMP-AMP synthase
C: Cyclic GMP-AMP synthase
E: Palindromic DNA18
F: Palindromic DNA18
I: Palindromic DNA18
J: Palindromic DNA18
hetero molecules


Theoretical massNumber of molelcules
Total (without water)108,86312
Polymers107,3396
Non-polymers1,5246
Water1,08160
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: HPLC of cGAS reaction using ITP+ATP as substrate shows a single dinucleotide peak. The retention time of the peak suggests it is a ATP-ITP.
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area10810 Å2
ΔGint-66 kcal/mol
Surface area40670 Å2
MethodPISA
Unit cell
Length a, b, c (Å)77.045, 98.534, 141.706
Angle α, β, γ (deg.)90.000, 90.000, 90.000
Int Tables number19
Space group name H-MP212121

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Components

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Protein / DNA chain , 2 types, 6 molecules ACEFIJ

#1: Protein Cyclic GMP-AMP synthase / cGAMP synthase / cGAS / m-cGAS / 2'3'-cGAMP synthase / Mab-21 domain-containing protein 1


Mass: 42640.254 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Details: Mouse Cyclic GMP-AMP synthase catalytic domain 147-507; There are three extra residues GTG at the N terminus (G from TEV site, TG from AgeI site).
Source: (gene. exp.) Mus musculus (house mouse) / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q8C6L5, cyclic GMP-AMP synthase
#2: DNA chain
Palindromic DNA18


Mass: 5514.603 Da / Num. of mol.: 4 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others)

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Non-polymers , 5 types, 66 molecules

#3: Chemical ChemComp-ATP / ADENOSINE-5'-TRIPHOSPHATE


Mass: 507.181 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C10H16N5O13P3 / Comment: ATP, energy-carrying molecule*YM
#4: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Mg
#5: Chemical ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Zn
#6: Chemical ChemComp-VWX / [[(2~{R},3~{R},4~{R},5~{R})-4-[[(2~{R},3~{S},4~{R},5~{R})-5-(6-aminopurin-9-yl)-3,4-bis(oxidanyl)oxolan-2-yl]methoxy-oxidanyl-phosphoryl]oxy-3-oxidanyl-5-(6-oxidanylidene-1~{H}-purin-9-yl)oxolan-2-yl]methoxy-oxidanyl-phosphoryl] phosphono hydrogen phosphate


Mass: 837.372 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C20H27N9O20P4 / Feature type: SUBJECT OF INVESTIGATION
#7: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 60 / Source method: isolated from a natural source / Formula: H2O

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Details

Has ligand of interestY

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 2.51 Å3/Da / Density % sol: 50.91 %
Crystal growTemperature: 277.15 K / Method: vapor diffusion, hanging drop / pH: 6.5
Details: 0.2 M ammonium acetate, 32% MPD, with 0.1 M Bis-Tris pH 6.5
PH range: 6.0-7.0 / Temp details: 4-degree Celsius in cold room

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Data collection

DiffractionMean temperature: 100 K / Ambient temp details: nitrogen gas stream / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: NSLS-II / Beamline: 17-ID-2 / Wavelength: 0.979307 Å
DetectorType: DECTRIS EIGER X 16M / Detector: PIXEL / Date: Aug 12, 2022
RadiationMonochromator: horizontal bounce Si(111) double crystal monochromator
Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.979307 Å / Relative weight: 1
ReflectionResolution: 2.44→80.9 Å / Num. obs: 34204 / % possible obs: 83.2 % / Redundancy: 1.9 % / CC1/2: 0.996 / Rmerge(I) obs: 0.067 / Rpim(I) all: 0.067 / Rrim(I) all: 0.095 / Net I/σ(I): 8.9 / Num. measured all: 64682
Reflection shell

Diffraction-ID: 1

Resolution (Å)Redundancy (%)Rmerge(I) obsNum. measured allNum. unique obsCC1/2Rpim(I) allRrim(I) allNet I/σ(I) obs% possible all
2.44-2.5320.60410185210.4620.6040.8541.312.4
9.12-80.91.70.01614768770.9990.0160.02223.799.3

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Processing

Software
NameVersionClassification
XDS20210205data reduction
Aimless0.7.9data scaling
MOLREP11.7.03phasing
Coot0.9.6model building
REFMAC5.8.0267refinement
PDB_EXTRACT3.27data extraction
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 7UUX

7uux


Resolution: 2.44→80.9 Å / Cor.coef. Fo:Fc: 0.927 / Cor.coef. Fo:Fc free: 0.914 / SU B: 28.597 / SU ML: 0.285 / Cross valid method: THROUGHOUT / ESU R: 0.887 / ESU R Free: 0.324 / Stereochemistry target values: MAXIMUM LIKELIHOOD
Details: U VALUES : WITH TLS ADDED HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS U VALUES : RESIDUAL ONLY
RfactorNum. reflection% reflectionSelection details
Rfree0.2586 1716 5 %RANDOM
Rwork0.2221 ---
obs0.2239 32438 83.27 %-
Solvent computationIon probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1 Å / Solvent model: MASK
Displacement parametersBiso max: 139.19 Å2 / Biso mean: 56.093 Å2 / Biso min: 23.44 Å2
Baniso -1Baniso -2Baniso -3
1--1.74 Å2-0 Å2-0 Å2
2---0.24 Å20 Å2
3---1.98 Å2
Refinement stepCycle: final / Resolution: 2.44→80.9 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms5701 1464 88 60 7313
Biso mean--61.58 38.46 -
Num. residues----761
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
X-RAY DIFFRACTIONr_bond_refined_d0.010.0167642
X-RAY DIFFRACTIONr_bond_other_d0.0010.026534
X-RAY DIFFRACTIONr_angle_refined_deg1.1641.71310496
X-RAY DIFFRACTIONr_angle_other_deg1.0682.7715190
X-RAY DIFFRACTIONr_dihedral_angle_1_deg6.045682
X-RAY DIFFRACTIONr_dihedral_angle_2_deg34.18722.524309
X-RAY DIFFRACTIONr_dihedral_angle_3_deg15.25151163
X-RAY DIFFRACTIONr_dihedral_angle_4_deg13.9841536
X-RAY DIFFRACTIONr_chiral_restr0.060.21079
X-RAY DIFFRACTIONr_gen_planes_refined0.0050.027220
X-RAY DIFFRACTIONr_gen_planes_other0.0010.021658
X-RAY DIFFRACTIONr_nbd_refined
X-RAY DIFFRACTIONr_nbd_other
X-RAY DIFFRACTIONr_nbtor_refined
X-RAY DIFFRACTIONr_nbtor_other
X-RAY DIFFRACTIONr_xyhbond_nbd_refined
X-RAY DIFFRACTIONr_xyhbond_nbd_other
X-RAY DIFFRACTIONr_metal_ion_refined
X-RAY DIFFRACTIONr_metal_ion_other
X-RAY DIFFRACTIONr_symmetry_vdw_refined
X-RAY DIFFRACTIONr_symmetry_vdw_other
X-RAY DIFFRACTIONr_symmetry_hbond_refined
X-RAY DIFFRACTIONr_symmetry_hbond_other
X-RAY DIFFRACTIONr_symmetry_metal_ion_refined
X-RAY DIFFRACTIONr_symmetry_metal_ion_other
X-RAY DIFFRACTIONr_rigid_bond_restr
X-RAY DIFFRACTIONr_sphericity_free
X-RAY DIFFRACTIONr_sphericity_bonded
LS refinement shellResolution: 2.44→2.501 Å / Rfactor Rfree error: 0
RfactorNum. reflection% reflection
Rfree0.396 9 -
Rwork0.293 196 -
obs--6.81 %
Refinement TLS params.

Method: refined / Refine-ID: X-RAY DIFFRACTION

IDL11 (°2)L12 (°2)L13 (°2)L22 (°2)L23 (°2)L33 (°2)S11 (Å °)S12 (Å °)S13 (Å °)S21 (Å °)S22 (Å °)S23 (Å °)S31 (Å °)S32 (Å °)S33 (Å °)T11 (Å2)T12 (Å2)T13 (Å2)T22 (Å2)T23 (Å2)T33 (Å2)Origin x (Å)Origin y (Å)Origin z (Å)
11.636-0.45410.36412.3257-0.07981.44030.0075-0.1259-0.22530.07480.0264-0.20110.18930.158-0.03390.08590.0094-0.00490.08840.020.06333.153-27.15420.105
21.8963-0.9627-0.34972.84150.61191.1897-0.0678-0.25610.16350.05510.06320.2948-0.1779-0.19070.00460.1110.0429-0.02420.1632-0.02050.0875.3638.22118.942
33.4041-0.293-1.0871.3101-1.90283.68050.0493-0.6330.08570.2570.2050.0398-0.2187-0.164-0.25430.2927-0.0896-0.09040.22570.07230.12349.701-23.1131.65
42.4382-0.515-2.02232.9783-0.34245.46290.1576-0.51740.03630.3342-0.11910.0486-0.4489-0.468-0.03860.1991-0.00150.0220.42080.00830.11219.069-23.60732.203
53.1501-1.80552.78214.07510.48924.34530.00780.17190.4123-0.1075-0.014-0.3009-0.16350.36390.00620.1215-0.08320.01710.1754-0.0580.299730.3778.51517.996
64.6724-0.51010.50464.6571-0.65363.61840.08030.50350.3556-0.2331-0.0043-0.4304-0.01670.6221-0.07610.2013-0.0066-0.05250.2497-0.09540.2130.016.7218.482
Refinement TLS group
IDRefine-IDRefine TLS-IDAuth asym-IDAuth seq-ID
1X-RAY DIFFRACTION1A149 - 506
2X-RAY DIFFRACTION2C149 - 506
3X-RAY DIFFRACTION3E1 - 18
4X-RAY DIFFRACTION4F1 - 18
5X-RAY DIFFRACTION5I1 - 18
6X-RAY DIFFRACTION6J1 - 18

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