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- EMDB-28545: Structure of SARS-CoV-2 Orf3a in plasma membrane-like environment... -
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Open data
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Basic information
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Title | Structure of SARS-CoV-2 Orf3a in plasma membrane-like environment, MSP1D1 nanodisc | |||||||||
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![]() | Membrane protein / SARS-CoV-2 / VIRAL PROTEIN | |||||||||
Function / homology | ![]() host cell lysosome / induction by virus of host reticulophagy / Maturation of protein 3a / SARS-CoV-2 modulates autophagy / inorganic cation transmembrane transport / voltage-gated calcium channel complex / host cell endoplasmic reticulum / SARS-CoV-2 targets host intracellular signalling and regulatory pathways / voltage-gated potassium channel complex / molecular function activator activity ...host cell lysosome / induction by virus of host reticulophagy / Maturation of protein 3a / SARS-CoV-2 modulates autophagy / inorganic cation transmembrane transport / voltage-gated calcium channel complex / host cell endoplasmic reticulum / SARS-CoV-2 targets host intracellular signalling and regulatory pathways / voltage-gated potassium channel complex / molecular function activator activity / cytoplasmic side of plasma membrane / protein complex oligomerization / monoatomic ion channel activity / host cell endosome / Translation of Structural Proteins / Virion Assembly and Release / Induction of Cell-Cell Fusion / Attachment and Entry / host cell endoplasmic reticulum membrane / SARS-CoV-2 activates/modulates innate and adaptive immune responses / host cell plasma membrane / virion membrane / extracellular region / identical protein binding / plasma membrane Similarity search - Function | |||||||||
Biological species | ![]() ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.4 Å | |||||||||
![]() | Miller AN / Houlihan PR / Matamala E / Cabezas-Bratesco D / Lee GY / Cristofori-Armstrong B / Dilan TL / Sanchez-Martinez S / Matthies D / Yan R ...Miller AN / Houlihan PR / Matamala E / Cabezas-Bratesco D / Lee GY / Cristofori-Armstrong B / Dilan TL / Sanchez-Martinez S / Matthies D / Yan R / Yu Z / Ren D / Brauchi SE / Clapham DE | |||||||||
Funding support | ![]()
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![]() | Journal: bioRxiv / Year: 2022 Title: The SARS-CoV-2 accessory protein Orf3a is not an ion channel, but does interact with trafficking proteins. Authors: Alexandria N Miller / Patrick R Houlihan / Ella Matamala / Deny Cabezas-Bratesco / Gi Young Lee / Ben Cristofori-Armstrong / Tanya L Dilan / Silvia Sanchez-Martinez / Doreen Matthies / Rui ...Authors: Alexandria N Miller / Patrick R Houlihan / Ella Matamala / Deny Cabezas-Bratesco / Gi Young Lee / Ben Cristofori-Armstrong / Tanya L Dilan / Silvia Sanchez-Martinez / Doreen Matthies / Rui Yan / Zhiheng Yu / Dejian Ren / Sebastian E Brauchi / David E Clapham Abstract: The severe acute respiratory syndrome associated coronavirus 2 (SARS-CoV-2) and SARS-CoV-1 accessory protein Orf3a colocalizes with markers of the plasma membrane, endocytic pathway, and Golgi ...The severe acute respiratory syndrome associated coronavirus 2 (SARS-CoV-2) and SARS-CoV-1 accessory protein Orf3a colocalizes with markers of the plasma membrane, endocytic pathway, and Golgi apparatus. Some reports have led to annotation of both Orf3a proteins as a viroporin. Here we show that neither SARS-CoV-2 nor SARS-CoV-1 form functional ion conducting pores and that the conductances measured are common contaminants in overexpression and with high levels of protein in reconstitution studies. Cryo-EM structures of both SARS-CoV-2 and SARS-CoV-1 Orf3a display a narrow constriction and the presence of a basic aqueous vestibule, which would not favor cation permeation. We observe enrichment of the late endosomal marker Rab7 upon SARS-CoV-2 Orf3a overexpression, and co-immunoprecipitation with VPS39. Interestingly, SARS-CoV-1 Orf3a does not cause the same cellular phenotype as SARS-CoV-2 Orf3a and does not interact with VPS39. To explain this difference, we find that a divergent, unstructured loop of SARS-CoV-2 Orf3a facilitates its binding with VPS39, a HOPS complex tethering protein involved in late endosome and autophagosome fusion with lysosomes. We suggest that the added loop enhances SARS-CoV-2 Orf3a ability to co-opt host cellular trafficking mechanisms for viral exit or host immune evasion. | |||||||||
History |
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Structure visualization
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 4.5 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 17.6 KB 17.6 KB | Display Display | ![]() |
Images | ![]() | 67 KB | ||
Filedesc metadata | ![]() | 5.9 KB | ||
Others | ![]() ![]() | 48.6 MB 48.6 MB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 8eqtMC ![]() 8eqjC ![]() 8eqsC ![]() 8equC M: atomic model generated by this map C: citing same article ( |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
EMDB pages | ![]() ![]() |
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Map
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Voxel size | X=Y=Z: 0.844 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Half map: #2
File | emd_28545_half_map_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #1
File | emd_28545_half_map_2.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
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Sample components
-Entire : Structure of SARS-CoV2 Orf3a in plasma membrane-like environment,...
Entire | Name: Structure of SARS-CoV2 Orf3a in plasma membrane-like environment, MSP1D1 nanodisc |
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Components |
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-Supramolecule #1: Structure of SARS-CoV2 Orf3a in plasma membrane-like environment,...
Supramolecule | Name: Structure of SARS-CoV2 Orf3a in plasma membrane-like environment, MSP1D1 nanodisc type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1 |
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Source (natural) | Organism: ![]() ![]() |
-Macromolecule #1: ORF3a protein
Macromolecule | Name: ORF3a protein / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 36.489445 KDa |
Recombinant expression | Organism: ![]() |
Sequence | String: MDLFMRIFTI GTVTLKQGEI KDATPSDFVR ATATIPIQAS LPFGWLIVGV ALLAVFQSAS KIITLKKRWQ LALSKGVHFV CNLLLLFVT VYSHLLLVAA GLEAPFLYLY ALVYFLQSIN FVRIIMRLWL CWKCRSKNPL LYDANYFLCW HTNCYDYCIP Y NSVTSSIV ...String: MDLFMRIFTI GTVTLKQGEI KDATPSDFVR ATATIPIQAS LPFGWLIVGV ALLAVFQSAS KIITLKKRWQ LALSKGVHFV CNLLLLFVT VYSHLLLVAA GLEAPFLYLY ALVYFLQSIN FVRIIMRLWL CWKCRSKNPL LYDANYFLCW HTNCYDYCIP Y NSVTSSIV ITSGDGTTSP ISEHDYQIGG YTEKWESGVK DCVVLHSYFT SDYYQLYSTQ LSTDTGVEHV TFFIYNKIVD EP EEHVQIH TIDGSSGVVN PVMEPIYDEP TTTTSVPLGG RGLEVLFQGP GSGQLVGSGG LEGGGGWSHP QFEKGGGSGG GSG GGSWSH PQFEK UniProtKB: ORF3a protein |
-Macromolecule #2: 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine
Macromolecule | Name: 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine / type: ligand / ID: 2 / Number of copies: 4 / Formula: PEE |
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Molecular weight | Theoretical: 744.034 Da |
Chemical component information | ![]() ChemComp-PEE: |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Buffer | pH: 7.5 |
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Vitrification | Cryogen name: ETHANE / Instrument: FEI VITROBOT MARK IV |
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Electron microscopy
Microscope | FEI TITAN KRIOS |
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Specialist optics | Energy filter - Name: GIF Bioquantum |
Image recording | Film or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 1 / Number real images: 15637 / Average electron dose: 50.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.8 µm |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Image processing
Startup model | Type of model: PDB ENTRY PDB model - PDB ID: |
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Final reconstruction | Number classes used: 1 / Applied symmetry - Point group: C2 (2 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 125625 |
Initial angle assignment | Type: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.0) |
Final angle assignment | Type: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1) |
-Atomic model buiding 1
Refinement | Protocol: AB INITIO MODEL |
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Output model | ![]() PDB-8eqt: |