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- PDB-8g10: Structure of Ternary Complex of cGAS with dsDNA and Bound ITP and GTP -

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Basic information

Entry
Database: PDB / ID: 8g10
TitleStructure of Ternary Complex of cGAS with dsDNA and Bound ITP and GTP
Components
  • Cyclic GMP-AMP synthase
  • Palindromic DNA18
KeywordsTRANSFERASE/DNA / IMMUNE SYSTEM/DNA / TRANSFERASE-DNA complex
Function / homology
Function and homology information


regulation of type I interferon production / cyclic GMP-AMP synthase / 2',3'-cyclic GMP-AMP synthase activity / paracrine signaling / poly-ADP-D-ribose modification-dependent protein binding / negative regulation of DNA repair / cGAS/STING signaling pathway / regulation of immunoglobulin production / regulation of T cell activation / pattern recognition receptor signaling pathway ...regulation of type I interferon production / cyclic GMP-AMP synthase / 2',3'-cyclic GMP-AMP synthase activity / paracrine signaling / poly-ADP-D-ribose modification-dependent protein binding / negative regulation of DNA repair / cGAS/STING signaling pathway / regulation of immunoglobulin production / regulation of T cell activation / pattern recognition receptor signaling pathway / negative regulation of cGAS/STING signaling pathway / cytoplasmic pattern recognition receptor signaling pathway / cellular response to exogenous dsRNA / cGMP-mediated signaling / regulation of immune response / positive regulation of type I interferon production / 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 / cAMP-mediated signaling / molecular condensate scaffold activity / determination of adult lifespan / positive regulation of cellular senescence / site of double-strand break / double-stranded DNA binding / defense response to virus / nuclear body / DNA repair / innate immune response / DNA damage response / chromatin binding / GTP binding / protein homodimerization activity / DNA binding / nucleoplasm / ATP binding / metal ion binding / nucleus / plasma membrane / cytosol / cytoplasm
Similarity search - Function
Mab-21-like, nucleotidyltransferase domain / Mab-21-like, HhH/H2TH-like domain / Mab-21 protein HhH/H2TH-like domain / Mab-21 protein nucleotidyltransferase domain / Mab-21-like / Mab-21
Similarity search - Domain/homology
GUANOSINE-5'-TRIPHOSPHATE / 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: 2.47 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: Nat Commun / Year: 2024
Title: The structural basis for 2'-5'/3'-5'-cGAMP synthesis by cGAS.
Authors: Wu, S. / Gabelli, S.B. / Sohn, J.
History
DepositionFeb 1, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0May 22, 2024Provider: repository / Type: Initial release
Revision 1.1May 29, 2024Group: Database references / Category: citation
Item: _citation.page_last / _citation.pdbx_database_id_PubMed / _citation.title

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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)109,60714
Polymers107,3356
Non-polymers2,2728
Water1,56787
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: isothermal titration calorimetry
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area13220 Å2
ΔGint-67 kcal/mol
Surface area41030 Å2
MethodPISA
Unit cell
Length a, b, c (Å)78.012, 98.659, 142.275
Angle α, β, γ (deg.)90.00, 90.00, 90.00
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: 42638.281 Da / Num. of mol.: 2 / Fragment: catalytic domain, residues 147-507 / Mutation: E211Q, D213N
Source method: isolated from a genetically manipulated source
Details: Mouse Cyclic GMP-AMP synthase catalytic domain 147-507; three extra residues at the N terminus (G from TEV site, TG from AgeI site). Flexible residues in the coordinate were deleted. Mutations:E211Q, D213N.
Source: (gene. exp.) Mus musculus (house mouse) / Gene: Cgas, Mb21d1 / Plasmid: nHMT mCAT WT QN
Details (production host): His*6-MBP-Tev-AgeI-mcGAS CAT, Kanamycin resistance
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 , 4 types, 95 molecules

#3: Chemical
ChemComp-GTP / GUANOSINE-5'-TRIPHOSPHATE / Guanosine triphosphate


Mass: 523.180 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C10H16N5O14P3 / Feature type: SUBJECT OF INVESTIGATION / Comment: GTP, 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: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 87 / 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: 2.55 Å3/Da / Density % sol: 51.77 %
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-1 / Wavelength: 0.92 Å
DetectorType: DECTRIS EIGER X 9M / Detector: PIXEL / Date: Sep 19, 2022
RadiationMonochromator: horizontal bounce Si(111) double crystal monochromator
Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.92 Å / Relative weight: 1
ReflectionResolution: 2.47→29.64 Å / Num. obs: 40157 / % possible obs: 99.7 % / Redundancy: 6.8 % / CC1/2: 0.999 / Rmerge(I) obs: 0.079 / Rpim(I) all: 0.033 / Rrim(I) all: 0.086 / Χ2: 1.01 / Net I/σ(I): 13.7 / Num. measured all: 271518
Reflection shellResolution: 2.47→2.57 Å / % possible obs: 97.8 % / Redundancy: 6.7 % / Rmerge(I) obs: 0.74 / Num. measured all: 29447 / Num. unique obs: 4379 / CC1/2: 0.802 / Rpim(I) all: 0.304 / Rrim(I) all: 0.801 / Χ2: 1.03 / Net I/σ(I) obs: 2.6

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Processing

Software
NameVersionClassification
XDS20210205data reduction
Aimless0.7.7data scaling
MOLREP11.7.03phasing
Coot0.9.6model building
PHENIX5.8.0403refinement
RefinementMethod to determine structure: MOLECULAR REPLACEMENT / Resolution: 2.47→29.64 Å / SU ML: 0.23 / Cross valid method: FREE R-VALUE / σ(F): 1.98 / Phase error: 23.41 / Stereochemistry target values: ML
RfactorNum. reflection% reflection
Rfree0.2272 2000 4.99 %
Rwork0.1936 --
obs0.1953 40055 99.63 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.1 Å / Solvent model: FLAT BULK SOLVENT MODEL
Refinement stepCycle: LAST / Resolution: 2.47→29.64 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms5766 1464 130 87 7447
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.0087662
X-RAY DIFFRACTIONf_angle_d1.15410639
X-RAY DIFFRACTIONf_dihedral_angle_d28.6651511
X-RAY DIFFRACTIONf_chiral_restr0.0521154
X-RAY DIFFRACTIONf_plane_restr0.0121073
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
2.47-2.530.28491360.25522584X-RAY DIFFRACTION96
2.53-2.60.26691400.2492685X-RAY DIFFRACTION100
2.6-2.670.2931430.25732695X-RAY DIFFRACTION100
2.67-2.760.27851390.23582656X-RAY DIFFRACTION100
2.76-2.860.28741420.22982712X-RAY DIFFRACTION100
2.86-2.970.23981410.23292689X-RAY DIFFRACTION100
2.97-3.110.29131440.24142721X-RAY DIFFRACTION100
3.11-3.270.23241410.22462701X-RAY DIFFRACTION100
3.27-3.480.25371430.19962717X-RAY DIFFRACTION100
3.48-3.740.22891430.19932721X-RAY DIFFRACTION100
3.75-4.120.23511440.18292739X-RAY DIFFRACTION100
4.12-4.710.19521450.15652750X-RAY DIFFRACTION100
4.71-5.930.19351460.17252796X-RAY DIFFRACTION100
5.93-29.640.19371530.16572889X-RAY DIFFRACTION100
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)
11.8825-0.5949-0.31882.7758-0.11241.51220.028-0.04450.1190.0845-0.02890.2632-0.1661-0.15180.00010.27340.0234-0.00870.3066-0.0440.32895.41327.09319.728
22.3619-1.09470.26363.0105-0.3761.6751-0.1174-0.2804-0.14420.10890.0826-0.24710.18310.12960.0010.33550.06550.05740.38590.04520.377933.595-7.9618.874
32.6087-0.43730.06781.68371.53931.19060.2123-0.83830.04830.57390.1326-0.0234-0.22080.26290.01050.5922-0.16170.06310.6968-0.02960.446928.99523.21831.54
41.9075-0.77450.83172.18140.55530.60380.3553-0.61480.07740.4553-0.13260.13180.5210.82770.00040.7417-0.0711-0.01980.7429-0.03780.438129.73623.78532.432
52.683-1.1848-1.33012.1883-0.23020.90330.10320.1338-0.5846-0.03050.16580.13530.4653-0.57790.00030.5975-0.1540.01280.58670.01920.67558.226-8.72817.901
62.9794-0.22290.1542.0142-0.88220.35070.19290.4293-0.3364-0.15630.1110.4688-0.1905-0.58810.00060.7069-0.02890.10140.6890.09620.62188.7-6.71618.683
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection detailsAuth asym-IDAuth seq-ID
1X-RAY DIFFRACTION1( CHAIN A AND ( RESID 145:507 OR RESID 604:604 ) )A145 - 507
2X-RAY DIFFRACTION1( CHAIN A AND ( RESID 145:507 OR RESID 604:604 ) )A604
3X-RAY DIFFRACTION2( CHAIN C AND ( RESID 149:507 OR RESID 604:604 ) )C149 - 507
4X-RAY DIFFRACTION2( CHAIN C AND ( RESID 149:507 OR RESID 604:604 ) )C604
5X-RAY DIFFRACTION3( CHAIN E AND RESID 1:18 )E1 - 18
6X-RAY DIFFRACTION4( CHAIN F AND RESID 1:18 )F1 - 18
7X-RAY DIFFRACTION5( CHAIN I AND RESID 1:18 )I1 - 18
8X-RAY DIFFRACTION6( CHAIN J AND RESID 1:18 )J1 - 18

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