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- PDB-5jc7: Crystal structure of chicken MDA5 with 5'p 24-mer dsRNA and ADP-M... -

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Basic information

Entry
Database: PDB / ID: 5jc7
TitleCrystal structure of chicken MDA5 with 5'p 24-mer dsRNA and ADP-Mg2+ at 2.75 A resolution.
Components
  • Melanoma differentiation associated protein-5
  • RNA (5'-R(P*GP*GP*AP*CP*GP*UP*CP*AP*UP*GP*CP*GP*CP*AP*UP*GP*AP*CP*GP*UP*CP*CP*CP*C)-3')
  • RNA (5'-R(P*GP*GP*GP*AP*CP*GP*UP*CP*AP*UP*GP*CP*GP*CP*AP*UP*GP*AP*CP*GP*UP*CP*CP*C)-3')
KeywordsIMMUNE SYSTEM / Innate immune pattern recognition receptor / RIG-I like helicase / dsRNA dependent ATPase / zinc-containing CTD domain
Function / homology
Function and homology information


Caspase-8 and -10 mediated induction of NF-kB / RLR (RIG-like receptor) mediated induction of IFN alpha/beta / TRAF mediated activation of IRF / Negative Regulation of MDA5 signaling / TRAF6 mediated NF-kB activation / defense response to virus / RNA helicase activity / hydrolase activity / RNA helicase / innate immune response ...Caspase-8 and -10 mediated induction of NF-kB / RLR (RIG-like receptor) mediated induction of IFN alpha/beta / TRAF mediated activation of IRF / Negative Regulation of MDA5 signaling / TRAF6 mediated NF-kB activation / defense response to virus / RNA helicase activity / hydrolase activity / RNA helicase / innate immune response / DNA binding / RNA binding / ATP binding / metal ion binding / cytoplasm
Similarity search - Function
phosphoenolpyruvate carboxylase, domain 3 - #30 / phosphoenolpyruvate carboxylase, domain 3 / RIG-I-like receptor, C-terminal regulatory domain / Metal Binding Protein, Guanine Nucleotide Exchange Factor; Chain A / RIG-I-like receptor, C-terminal / RIG-I receptor C-terminal domain / RIG-I-like receptor, C-terminal regulatory domain / RIG-I-like receptor, C-terminal domain superfamily / : / C-terminal domain of RIG-I ...phosphoenolpyruvate carboxylase, domain 3 - #30 / phosphoenolpyruvate carboxylase, domain 3 / RIG-I-like receptor, C-terminal regulatory domain / Metal Binding Protein, Guanine Nucleotide Exchange Factor; Chain A / RIG-I-like receptor, C-terminal / RIG-I receptor C-terminal domain / RIG-I-like receptor, C-terminal regulatory domain / RIG-I-like receptor, C-terminal domain superfamily / : / C-terminal domain of RIG-I / RIG-I-like receptor (RLR) C-terminal regulatory (CTR) domain profile. / Caspase recruitment domain / Caspase recruitment domain / Helicase/UvrB, N-terminal / Type III restriction enzyme, res subunit / Death-like domain superfamily / Beta Complex / Helicase conserved C-terminal domain / helicase superfamily c-terminal domain / Superfamilies 1 and 2 helicase C-terminal domain profile. / Superfamilies 1 and 2 helicase ATP-binding type-1 domain profile. / DEAD-like helicases superfamily / Helicase, C-terminal / Helicase superfamily 1/2, ATP-binding domain / Armadillo-type fold / P-loop containing nucleotide triphosphate hydrolases / Up-down Bundle / Rossmann fold / P-loop containing nucleoside triphosphate hydrolase / 3-Layer(aba) Sandwich / Mainly Beta / Mainly Alpha / Alpha Beta
Similarity search - Domain/homology
ADENOSINE-5'-DIPHOSPHATE / RNA / RNA (> 10) / RNA helicase
Similarity search - Component
Biological speciesGallus gallus (chicken)
synthetic construct (others)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.75 Å
AuthorsCusack, S. / Uchikawa, E.
Funding support France, 1items
OrganizationGrant numberCountry
European Research Council322586 France
CitationJournal: Mol.Cell / Year: 2016
Title: Structural Analysis of dsRNA Binding to Anti-viral Pattern Recognition Receptors LGP2 and MDA5.
Authors: Uchikawa, E. / Lethier, M. / Malet, H. / Brunel, J. / Gerlier, D. / Cusack, S.
History
DepositionApr 14, 2016Deposition site: RCSB / Processing site: PDBE
Revision 1.0Jun 1, 2016Provider: repository / Type: Initial release
Revision 1.1Feb 20, 2019Group: Author supporting evidence / Data collection / Source and taxonomy
Category: pdbx_audit_support / pdbx_entity_src_syn
Item: _pdbx_audit_support.funding_organization / _pdbx_entity_src_syn.ncbi_taxonomy_id / _pdbx_entity_src_syn.organism_scientific
Revision 1.2Jan 10, 2024Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model / pdbx_struct_conn_angle / struct_conn / struct_ncs_dom_lim
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _pdbx_struct_conn_angle.ptnr1_auth_asym_id / _pdbx_struct_conn_angle.ptnr1_auth_comp_id / _pdbx_struct_conn_angle.ptnr1_auth_seq_id / _pdbx_struct_conn_angle.ptnr1_label_asym_id / _pdbx_struct_conn_angle.ptnr1_label_atom_id / _pdbx_struct_conn_angle.ptnr1_label_comp_id / _pdbx_struct_conn_angle.ptnr1_label_seq_id / _pdbx_struct_conn_angle.ptnr2_auth_asym_id / _pdbx_struct_conn_angle.ptnr2_auth_comp_id / _pdbx_struct_conn_angle.ptnr2_auth_seq_id / _pdbx_struct_conn_angle.ptnr2_label_asym_id / _pdbx_struct_conn_angle.ptnr2_label_atom_id / _pdbx_struct_conn_angle.ptnr2_label_comp_id / _pdbx_struct_conn_angle.ptnr3_auth_asym_id / _pdbx_struct_conn_angle.ptnr3_auth_comp_id / _pdbx_struct_conn_angle.ptnr3_auth_seq_id / _pdbx_struct_conn_angle.ptnr3_label_asym_id / _pdbx_struct_conn_angle.ptnr3_label_atom_id / _pdbx_struct_conn_angle.ptnr3_label_comp_id / _pdbx_struct_conn_angle.ptnr3_label_seq_id / _pdbx_struct_conn_angle.value / _struct_conn.pdbx_dist_value / _struct_conn.ptnr1_auth_asym_id / _struct_conn.ptnr1_auth_comp_id / _struct_conn.ptnr1_auth_seq_id / _struct_conn.ptnr1_label_asym_id / _struct_conn.ptnr1_label_atom_id / _struct_conn.ptnr1_label_comp_id / _struct_conn.ptnr1_label_seq_id / _struct_conn.ptnr2_auth_asym_id / _struct_conn.ptnr2_auth_comp_id / _struct_conn.ptnr2_auth_seq_id / _struct_conn.ptnr2_label_asym_id / _struct_conn.ptnr2_label_atom_id / _struct_conn.ptnr2_label_comp_id / _struct_ncs_dom_lim.beg_auth_comp_id / _struct_ncs_dom_lim.beg_label_asym_id / _struct_ncs_dom_lim.beg_label_comp_id / _struct_ncs_dom_lim.beg_label_seq_id / _struct_ncs_dom_lim.end_auth_comp_id / _struct_ncs_dom_lim.end_label_asym_id / _struct_ncs_dom_lim.end_label_comp_id / _struct_ncs_dom_lim.end_label_seq_id

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Melanoma differentiation associated protein-5
B: Melanoma differentiation associated protein-5
X: RNA (5'-R(P*GP*GP*AP*CP*GP*UP*CP*AP*UP*GP*CP*GP*CP*AP*UP*GP*AP*CP*GP*UP*CP*CP*CP*C)-3')
Y: RNA (5'-R(P*GP*GP*GP*AP*CP*GP*UP*CP*AP*UP*GP*CP*GP*CP*AP*UP*GP*AP*CP*GP*UP*CP*CP*C)-3')
hetero molecules


Theoretical massNumber of molelcules
Total (without water)180,07210
Polymers179,0384
Non-polymers1,0346
Water00
1


  • Idetical with deposited unit
  • defined by software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area13530 Å2
ΔGint-137 kcal/mol
Surface area63970 Å2
MethodPISA
Unit cell
Length a, b, c (Å)99.750, 133.400, 138.440
Angle α, β, γ (deg.)90.00, 90.00, 90.00
Int Tables number19
Space group name H-MP212121
Noncrystallographic symmetry (NCS)NCS domain:
IDEns-IDDetails
11A
21B
12X
22Y

NCS domain segments:

Component-ID: _ / Refine code: _

Dom-IDEns-IDBeg auth comp-IDBeg label comp-IDEnd auth comp-IDEnd label comp-IDAuth asym-IDLabel asym-IDAuth seq-IDLabel seq-ID
11ASPASPPHEPHEAA298 - 9895 - 696
21ASPASPPHEPHEBB298 - 9895 - 696
12GGCCXC1 - 222 - 23
22GGCCYD3 - 242 - 23

NCS ensembles :
ID
1
2

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Components

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Protein , 1 types, 2 molecules AB

#1: Protein Melanoma differentiation associated protein-5


Mass: 81667.008 Da / Num. of mol.: 2
Mutation: N-terminal deletion of 1-297; GAMG from tag at N-terminus; C-terminal deletion of 995-1001; Point mutation E436Q.
Source method: isolated from a genetically manipulated source
Details: Chicken MDA5 helicase-CTD domains / Source: (gene. exp.) Gallus gallus (chicken) / Gene: MDA5 / Production host: Escherichia coli BL21(DE3) (bacteria) / Variant (production host): Rosetta 2 / References: UniProt: D9N195

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RNA chain , 2 types, 2 molecules XY

#2: RNA chain RNA (5'-R(P*GP*GP*AP*CP*GP*UP*CP*AP*UP*GP*CP*GP*CP*AP*UP*GP*AP*CP*GP*UP*CP*CP*CP*C)-3')


Mass: 8004.811 Da / Num. of mol.: 1 / Source method: obtained synthetically / Details: Synthetic dsRNA using T7 polymerase / Source: (synth.) synthetic construct (others)
#3: RNA chain RNA (5'-R(P*GP*GP*GP*AP*CP*GP*UP*CP*AP*UP*GP*CP*GP*CP*AP*UP*GP*AP*CP*GP*UP*CP*CP*C)-3')


Mass: 7699.629 Da / Num. of mol.: 1 / Source method: obtained synthetically / Details: Synthetic dsRNA using T7 polymerase / Source: (synth.) synthetic construct (others)

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Non-polymers , 3 types, 6 molecules

#4: Chemical ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Zn
#5: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Mg
#6: Chemical ChemComp-ADP / ADENOSINE-5'-DIPHOSPHATE


Mass: 427.201 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C10H15N5O10P2 / Comment: ADP, energy-carrying molecule*YM

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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: 2.57 Å3/Da / Density % sol: 52.18 %
Crystal growTemperature: 293 K / Method: vapor diffusion, sitting drop / pH: 7.5
Details: Directly after size exclusion chromatography chMDA5 deltaCARD-Q was mixed with 24-mer dsRNA in a 0.5:1 molar ratio and incubated for 30 minutes on ice. The complexes were concentrated to ...Details: Directly after size exclusion chromatography chMDA5 deltaCARD-Q was mixed with 24-mer dsRNA in a 0.5:1 molar ratio and incubated for 30 minutes on ice. The complexes were concentrated to around 10 mg/ml and 2 mM AMPPNP and 200 mM NDSB211 (Dimethyl(2-hydroxyethyl)ammonium propane sulfonate) were added. chMDA5 delatCARD-Q:24bp dsRNA:AMPPNP was mixed with reservoir buffer (0.1 mM Hepes pH 7.5, 11-12% PEG 3350, 2-4% sucrose) in a 2:1 ratio. Crystals grew in three days at 20 degree and were harvested in cryo-protecting solution (0.1 mM Hepes pH 7.5, 25% 3350, 10% ethylene glycol) before flash freezing with liquid nitrogen

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: ESRF / Beamline: ID29 / Wavelength: 0.9724 Å
DetectorType: DECTRIS PILATUS3 6M / Detector: PIXEL / Date: Jul 25, 2013
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.9724 Å / Relative weight: 1
ReflectionResolution: 2.75→50 Å / Num. obs: 48273 / % possible obs: 99.1 % / Redundancy: 4.66 % / Rmerge(I) obs: 0.168 / Net I/σ(I): 6.58
Reflection shellResolution: 2.75→2.82 Å / Redundancy: 4.89 % / Rmerge(I) obs: 0.809 / Mean I/σ(I) obs: 1.52 / % possible all: 99.2

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Processing

Software
NameVersionClassification
REFMAC5.8.0131refinement
XDSdata reduction
XSCALEdata scaling
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 5JC3

5jc3


Resolution: 2.75→50 Å / Cor.coef. Fo:Fc: 0.919 / Cor.coef. Fo:Fc free: 0.898 / SU B: 34.451 / SU ML: 0.59 / Cross valid method: THROUGHOUT / ESU R Free: 0.455 / Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS
RfactorNum. reflection% reflectionSelection details
Rfree0.32199 2433 5 %RANDOM
Rwork0.29064 ---
obs0.29224 45840 99.08 %-
Solvent computationIon probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.2 Å
Displacement parametersBiso mean: 69.662 Å2
Baniso -1Baniso -2Baniso -3
1-7.4 Å20 Å20 Å2
2--2.21 Å2-0 Å2
3----9.61 Å2
Refinement stepCycle: 1 / Resolution: 2.75→50 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms10537 1021 58 0 11616
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
X-RAY DIFFRACTIONr_bond_refined_d0.0080.01911915
X-RAY DIFFRACTIONr_bond_other_d0.0020.0211099
X-RAY DIFFRACTIONr_angle_refined_deg1.2121.89416230
X-RAY DIFFRACTIONr_angle_other_deg0.9413.00125608
X-RAY DIFFRACTIONr_dihedral_angle_1_deg6.27951274
X-RAY DIFFRACTIONr_dihedral_angle_2_deg36.29924.229532
X-RAY DIFFRACTIONr_dihedral_angle_3_deg16.126152138
X-RAY DIFFRACTIONr_dihedral_angle_4_deg14.3861586
X-RAY DIFFRACTIONr_chiral_restr0.0660.21771
X-RAY DIFFRACTIONr_gen_planes_refined0.0040.0212467
X-RAY DIFFRACTIONr_gen_planes_other0.0020.022718
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_mcbond_it2.9726.9425138
X-RAY DIFFRACTIONr_mcbond_other2.976.9415137
X-RAY DIFFRACTIONr_mcangle_it5.07210.3946398
X-RAY DIFFRACTIONr_mcangle_other5.07210.3956399
X-RAY DIFFRACTIONr_scbond_it2.4096.9876777
X-RAY DIFFRACTIONr_scbond_other2.4096.9866776
X-RAY DIFFRACTIONr_scangle_it
X-RAY DIFFRACTIONr_scangle_other4.22410.3499832
X-RAY DIFFRACTIONr_long_range_B_refined7.32253.10612984
X-RAY DIFFRACTIONr_long_range_B_other7.32253.11112985
X-RAY DIFFRACTIONr_rigid_bond_restr
X-RAY DIFFRACTIONr_sphericity_free
X-RAY DIFFRACTIONr_sphericity_bonded
Refine LS restraints NCS

Refine-ID: X-RAY DIFFRACTION / Type: interatomic distance / Rms dev position: 0.02 Å / Weight position: 0.05

Ens-IDDom-IDAuth asym-IDNumber
11A84834
12B84834
21X3470
22Y3470
LS refinement shellResolution: 2.75→2.821 Å / Total num. of bins used: 20
RfactorNum. reflection% reflection
Rfree0.492 176 -
Rwork0.487 3359 -
obs--99.13 %

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