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- PDB-8ctk: Cryo-EM structure of SARS-CoV-2 M protein in a lipid nanodisc -

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Basic information

Entry
Database: PDB / ID: 8ctk
TitleCryo-EM structure of SARS-CoV-2 M protein in a lipid nanodisc
ComponentsMembrane protein
KeywordsMEMBRANE PROTEIN / SARS-COV-2 / CORONAVIRUS / VIRAL PROTEIN / CAPSID PROTEIN
Function / homology
Function and homology information


Maturation of protein M / SARS-CoV-2 modulates autophagy / cytoplasmic capsid assembly / endoplasmic reticulum-Golgi intermediate compartment / CD28 dependent PI3K/Akt signaling / host cell Golgi membrane / SARS-CoV-2 targets host intracellular signalling and regulatory pathways / protein sequestering activity / symbiont-mediated suppression of host cytoplasmic pattern recognition receptor signaling pathway via inhibition of MAVS activity / VEGFR2 mediated vascular permeability ...Maturation of protein M / SARS-CoV-2 modulates autophagy / cytoplasmic capsid assembly / endoplasmic reticulum-Golgi intermediate compartment / CD28 dependent PI3K/Akt signaling / host cell Golgi membrane / SARS-CoV-2 targets host intracellular signalling and regulatory pathways / protein sequestering activity / symbiont-mediated suppression of host cytoplasmic pattern recognition receptor signaling pathway via inhibition of MAVS activity / VEGFR2 mediated vascular permeability / PIP3 activates AKT signaling / TRAF3-dependent IRF activation pathway / Translation of Structural Proteins / Virion Assembly and Release / Induction of Cell-Cell Fusion / structural constituent of virion / Attachment and Entry / virus-mediated perturbation of host defense response / viral envelope / SARS-CoV-2 activates/modulates innate and adaptive immune responses / virion membrane / identical protein binding / plasma membrane
Similarity search - Function
M matrix/glycoprotein, SARS-CoV-like / M matrix/glycoprotein, coronavirus / Coronavirus M matrix/glycoprotein / Coronavirus membrane (Cov-M) protein profile.
Similarity search - Domain/homology
Biological speciesSevere acute respiratory syndrome coronavirus 2
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.52 Å
AuthorsDolan, K.A. / Brohawn, S.G.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM123496 United States
Citation
Journal: Elife / Year: 2022
Title: Structure of SARS-CoV-2 M protein in lipid nanodiscs.
Authors: Kimberly A Dolan / Mandira Dutta / David M Kern / Abhay Kotecha / Gregory A Voth / Stephen G Brohawn /
Abstract: SARS-CoV-2 encodes four structural proteins incorporated into virions, spike (S), envelope (E), nucleocapsid (N), and membrane (M). M plays an essential role in viral assembly by organizing other ...SARS-CoV-2 encodes four structural proteins incorporated into virions, spike (S), envelope (E), nucleocapsid (N), and membrane (M). M plays an essential role in viral assembly by organizing other structural proteins through physical interactions and directing them to sites of viral budding. As the most abundant protein in the viral envelope and a target of patient antibodies, M is a compelling target for vaccines and therapeutics. Still, the structure of M and molecular basis for its role in virion formation are unknown. Here, we present the cryo-EM structure of SARS-CoV-2 M in lipid nanodiscs to 3.5 Å resolution. M forms a 50 kDa homodimer that is structurally related to the SARS-CoV-2 ORF3a viroporin, suggesting a shared ancestral origin. Structural comparisons reveal how intersubunit gaps create a small, enclosed pocket in M and large open cavity in ORF3a, consistent with a structural role and ion channel activity, respectively. M displays a strikingly electropositive cytosolic surface that may be important for interactions with N, S, and viral RNA. Molecular dynamics simulations show a high degree of structural rigidity in a simple lipid bilayer and support a role for M homodimers in scaffolding viral assembly. Together, these results provide insight into roles for M in coronavirus assembly and structure.
#1: Journal: Acta Crystallogr., Sect. D: Biol. Crystallogr. / Year: 2018
Title: Real-space refinement in PHENIX for cryo-EM and crystallography
Authors: Afonine, P.V.
History
DepositionMay 15, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jun 22, 2022Provider: repository / Type: Initial release
Revision 1.1Nov 2, 2022Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year
Revision 1.2Jun 12, 2024Group: Data collection / Category: chem_comp_atom / chem_comp_bond

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Membrane protein
B: Membrane protein


Theoretical massNumber of molelcules
Total (without water)52,3812
Polymers52,3812
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551
Noncrystallographic symmetry (NCS)NCS domain:
IDEns-IDDetails
d_1ens_1chain "B"
d_2ens_1chain "A"

NCS domain segments:
Dom-IDComponent-IDEns-IDBeg label comp-IDEnd label comp-IDLabel asym-IDLabel seq-ID
d_11ens_1LEUTYRB1 - 188
d_21ens_1LEUTYRA1 - 188

NCS oper: (Code: givenMatrix: (-0.999992069585, -0.00208585001278, 0.003392638651), (0.00207034571065, -0.999987427585, -0.00456709312518), (0.00340212226859, -0.00456003297136, 0.999983815701)Vector: ...NCS oper: (Code: given
Matrix: (-0.999992069585, -0.00208585001278, 0.003392638651), (0.00207034571065, -0.999987427585, -0.00456709312518), (0.00340212226859, -0.00456003297136, 0.999983815701)
Vector: 209.132617949, 209.670532741, 0.0551990723641)

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Components

#1: Protein Membrane protein / M / E1 glycoprotein / Matrix glycoprotein / Membrane glycoprotein


Mass: 26190.693 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Severe acute respiratory syndrome coronavirus 2
Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P0DTC5

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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: M protein / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightValue: 0.052 MDa / Experimental value: NO
Source (natural)Organism: Severe acute respiratory syndrome coronavirus 2
Source (recombinant)Organism: Spodoptera frugiperda (fall armyworm)
Buffer solutionpH: 7.4
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMHEPESC8H18N2O4S1
2150 mMpotassium chlorideKCl1
SpecimenConc.: 1.3 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K / Details: blot force 1, wait time 5s, blot time 3s

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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 FIELD / Nominal defocus max: 1200 nm / Nominal defocus min: 500 nm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: FEI FALCON IV (4k x 4k)

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Processing

Software
NameVersionClassification
phenix.real_space_refine1.20.1_4487refinement
PHENIX1.20.1_4487refinement
EM software
IDNameVersionCategory
4CTFFINDCTF correction
11cryoSPARC3.1final Euler assignment
13cryoSPARC3.13D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C2 (2 fold cyclic)
3D reconstructionResolution: 3.52 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 64966 / Symmetry type: POINT
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 42.42 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00323126
ELECTRON MICROSCOPYf_angle_d0.68224254
ELECTRON MICROSCOPYf_chiral_restr0.0468486
ELECTRON MICROSCOPYf_plane_restr0.0047512
ELECTRON MICROSCOPYf_dihedral_angle_d3.9848420
Refine LS restraints NCSType: NCS constraints / Rms dev position: 8.62962799761E-12 Å

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