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- PDB-6nk6: Electron Cryo-Microscopy Of Chikungunya VLP in complex with mouse... -

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Basic information

Entry
Database: PDB / ID: 6nk6
TitleElectron Cryo-Microscopy Of Chikungunya VLP in complex with mouse Mxra8 receptor
Components
  • Capsid proteinCapsid
  • E1 glycoprotein
  • E2 glycoprotein
  • Matrix remodeling-associated protein 8
KeywordsVIRUS LIKE PARTICLE/SIGNALING PROTEIN / Chikungunya / virus-like particle / viral receptor / Mxra8 / Structural Genomics / Center for Structural Genomics of Infectious Diseases / CSGID / VIRUS LIKE PARTICLE-SIGNALING PROTEIN complex
Function / homology
Function and homology information


Regulation of Insulin-like Growth Factor (IGF) transport and uptake by Insulin-like Growth Factor Binding Proteins (IGFBPs) / Post-translational protein phosphorylation / establishment of glial blood-brain barrier / ec:3.4.21.90: / T=4 icosahedral viral capsid / ciliary membrane / bicellular tight junction / fusion of virus membrane with host endosome membrane / host cell cytoplasm / cell adhesion ...Regulation of Insulin-like Growth Factor (IGF) transport and uptake by Insulin-like Growth Factor Binding Proteins (IGFBPs) / Post-translational protein phosphorylation / establishment of glial blood-brain barrier / ec:3.4.21.90: / T=4 icosahedral viral capsid / ciliary membrane / bicellular tight junction / fusion of virus membrane with host endosome membrane / host cell cytoplasm / cell adhesion / host cell nucleus / virion attachment to host cell / host cell plasma membrane / virion membrane / serine-type endopeptidase activity / structural molecule activity / cell surface / RNA binding / integral component of membrane / nucleus / cytoplasm
Alphavirus E1 glycoprotein / Flaviviral glycoprotein E, dimerisation domain / Immunoglobulin-like domain / Immunoglobulin subtype / Immunoglobulin-like fold / Alphavirus E3 spike glycoprotein / Alphavirus E2 glycoprotein / Peptidase S3, togavirin / Flavivirus/Alphavirus glycoprotein, immunoglobulin-like domain superfamily / Immunoglobulin E-set ...Alphavirus E1 glycoprotein / Flaviviral glycoprotein E, dimerisation domain / Immunoglobulin-like domain / Immunoglobulin subtype / Immunoglobulin-like fold / Alphavirus E3 spike glycoprotein / Alphavirus E2 glycoprotein / Peptidase S3, togavirin / Flavivirus/Alphavirus glycoprotein, immunoglobulin-like domain superfamily / Immunoglobulin E-set / Immunoglobulin-like domain superfamily / Flavivirus glycoprotein, central and dimerisation domain superfamily / Alphavirus E2 glycoprotein, domain A / Immunoglobulin V-set domain / Alphavirus E2 glycoprotein, domain B / Alphavirus E2 glycoprotein, domain C / Alphavirus E2 glycoprotein / Alphavirus core protein / Alphavirus E3 glycoprotein / Alphavirus E1 glycoprotein / Immunoglobulin V-set domain / Ig-like domain profile. / Alphavirus core protein (CP) domain profile. / Peptidase S1, PA clan
Structural polyprotein / Matrix remodeling-associated protein 8
Biological speciesMus musculus (house mouse)
Chikungunya virus strain Senegal 37997
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.06 Å
AuthorsBasore, K. / Kim, A.S. / Nelson, C.A. / Fremont, D.H. / Center for Structural Genomics of Infectious Diseases (CSGID)
Funding supportUnited States , 1件
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious DiseasesHHSN272201700060CUnited States
CitationJournal: Cell / Year: 2019
Title: Cryo-EM Structure of Chikungunya Virus in Complex with the Mxra8 Receptor.
Authors: Katherine Basore / Arthur S Kim / Christopher A Nelson / Rong Zhang / Brittany K Smith / Carla Uranga / Lo Vang / Ming Cheng / Michael L Gross / Jonathan Smith / Michael S Diamond / Daved H Fremont /
Abstract: Mxra8 is a receptor for multiple arthritogenic alphaviruses that cause debilitating acute and chronic musculoskeletal disease in humans. Herein, we present a 2.2 Å resolution X-ray crystal ...Mxra8 is a receptor for multiple arthritogenic alphaviruses that cause debilitating acute and chronic musculoskeletal disease in humans. Herein, we present a 2.2 Å resolution X-ray crystal structure of Mxra8 and 4 to 5 Å resolution cryo-electron microscopy reconstructions of Mxra8 bound to chikungunya (CHIKV) virus-like particles and infectious virus. The Mxra8 ectodomain contains two strand-swapped Ig-like domains oriented in a unique disulfide-linked head-to-head arrangement. Mxra8 binds by wedging into a cleft created by two adjacent CHIKV E2-E1 heterodimers in one trimeric spike and engaging a neighboring spike. Two binding modes are observed with the fully mature VLP, with one Mxra8 binding with unique contacts. Only the high-affinity binding mode was observed in the complex with infectious CHIKV, as viral maturation and E3 occupancy appear to influence receptor binding-site usage. Our studies provide insight into how Mxra8 binds CHIKV and creates a path for developing alphavirus entry inhibitors.
Validation Report
SummaryFull reportAbout validation report
DateDeposition: Jan 4, 2019 / Release: May 22, 2019
RevisionDateData content typeGroupCategoryItemProviderType
1.0May 22, 2019Structure modelrepositoryInitial release
1.1May 29, 2019Structure modelData collection / Database referencescitation / citation_author_citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID / _citation_author.name
1.2Jun 26, 2019Structure modelData collection / Database referencescitation_citation.journal_volume / _citation.page_first

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Structure visualization

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  • Biological unit as complete icosahedral assembly
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  • Biological unit as icosahedral pentamer
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  • Biological unit as icosahedral 23 hexamer
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  • Deposited structure unit
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  • Simplified surface model + fitted atomic model
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  • Superimposition on EM map
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Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
M: Matrix remodeling-associated protein 8
N: Matrix remodeling-associated protein 8
O: Matrix remodeling-associated protein 8
P: Matrix remodeling-associated protein 8
A: E1 glycoprotein
E: E2 glycoprotein
I: Capsid protein
B: E1 glycoprotein
F: E2 glycoprotein
J: Capsid protein
C: E1 glycoprotein
G: E2 glycoprotein
K: Capsid protein
D: E1 glycoprotein
H: E2 glycoprotein
L: Capsid protein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)568,58728
Polymers565,93316
Non-polymers2,65412
Water0
1
M: Matrix remodeling-associated protein 8
N: Matrix remodeling-associated protein 8
O: Matrix remodeling-associated protein 8
P: Matrix remodeling-associated protein 8
A: E1 glycoprotein
E: E2 glycoprotein
I: Capsid protein
B: E1 glycoprotein
F: E2 glycoprotein
J: Capsid protein
C: E1 glycoprotein
G: E2 glycoprotein
K: Capsid protein
D: E1 glycoprotein
H: E2 glycoprotein
L: Capsid protein
hetero molecules
x 60


Theoretical massNumber of molelcules
Total (without water)34,115,2341680
Polymers33,955,964960
Non-polymers159,270720
Water0
TypeNameSymmetry operationNumber
point symmetry operation60
2


  • Idetical with deposited unit in distinct coordinate
  • icosahedral asymmetric unit
TypeNameSymmetry operationNumber
point symmetry operation1
3
M: Matrix remodeling-associated protein 8
N: Matrix remodeling-associated protein 8
O: Matrix remodeling-associated protein 8
P: Matrix remodeling-associated protein 8
A: E1 glycoprotein
E: E2 glycoprotein
I: Capsid protein
B: E1 glycoprotein
F: E2 glycoprotein
J: Capsid protein
C: E1 glycoprotein
G: E2 glycoprotein
K: Capsid protein
D: E1 glycoprotein
H: E2 glycoprotein
L: Capsid protein
hetero molecules
x 5


  • icosahedral pentamer
  • 2.84 MDa, 80 polymers
Theoretical massNumber of molelcules
Total (without water)2,842,936140
Polymers2,829,66480
Non-polymers13,27260
Water0
TypeNameSymmetry operationNumber
point symmetry operation5
4
M: Matrix remodeling-associated protein 8
N: Matrix remodeling-associated protein 8
O: Matrix remodeling-associated protein 8
P: Matrix remodeling-associated protein 8
A: E1 glycoprotein
E: E2 glycoprotein
I: Capsid protein
B: E1 glycoprotein
F: E2 glycoprotein
J: Capsid protein
C: E1 glycoprotein
G: E2 glycoprotein
K: Capsid protein
D: E1 glycoprotein
H: E2 glycoprotein
L: Capsid protein
hetero molecules
x 6


  • icosahedral 23 hexamer
  • 3.41 MDa, 96 polymers
Theoretical massNumber of molelcules
Total (without water)3,411,523168
Polymers3,395,59696
Non-polymers15,92772
Water0
TypeNameSymmetry operationNumber
point symmetry operation6
5


  • Idetical with deposited unit in distinct coordinate
  • icosahedral asymmetric unit, std point frame
TypeNameSymmetry operationNumber
transform to point frame1
SymmetryPoint symmetry: (Schoenflies symbol: I (icosahedral))

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Components

#1: Protein/peptide
Matrix remodeling-associated protein 8 / Adipocyte-specific protein 3 / Dual Ig domain-containing cell adhesion molecule / DICAM / Limitrin / Mxra8 receptor


Mass: 30819.457 Da / Num. of mol.: 4 / Fragment: ectodomain (UNP residues 39-291)
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mus musculus (house mouse) / Gene: Mxra8, Asp3, Dicam / Cell line (production host): HEK293F / Production host: Homo sapiens (human) / References: UniProt: Q9DBV4
#2: Protein/peptide
E1 glycoprotein


Mass: 47346.715 Da / Num. of mol.: 4 / Fragment: UNP residues 810-1248
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Chikungunya virus strain Senegal 37997 / Strain: 37997 / Cell line (production host): HEK293F / Production host: Homo sapiens (human) / References: UniProt: Q5XXP3
#3: Protein/peptide
E2 glycoprotein


Mass: 46858.312 Da / Num. of mol.: 4 / Fragment: UNP residues 330-748
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Chikungunya virus strain Senegal 37997 / Strain: 37997 / Cell line (production host): HEK293F / Production host: Homo sapiens (human) / References: UniProt: Q5XXP3
#4: Protein/peptide
Capsid protein / Capsid


Mass: 16458.701 Da / Num. of mol.: 4 / Fragment: UNP residues 111-261
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Chikungunya virus strain Senegal 37997 / Strain: 37997 / Cell line (production host): HEK293F / Production host: Homo sapiens (human) / References: UniProt: Q5XXP3
#5: Chemical
ChemComp-NAG / N-ACETYL-D-GLUCOSAMINE


Mass: 221.208 Da / Num. of mol.: 12
Source method: isolated from a genetically manipulated source
Formula: C8H15NO6 / N-Acetylglucosamine

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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

Component

Type: COMPLEX

IDNameDetailsEntity IDParent-IDSource
1Chikungunya virus like particle with Mxra8VLP Produced by PaxVax Corporation, Redwood CA, USA. Mouse Mxra8 produced in HEK293 cells.1, 2, 3, 40MULTIPLE SOURCES
2Chikungunya virus like particle2, 3, 41RECOMBINANT
3Mxra8 receptor11RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-IDStrain
12Chikungunya virus (strain 37997)37109537997
23Mus musculus (house mouse)10090
Source (recombinant)

Ncbi tax-ID: 9606 / Organism: Homo sapiens (human) / Strain: HEK293F

IDEntity assembly-ID
12
23
Details of virusEmpty: YES / Enveloped: YES / Isolate: STRAIN / Type: VIRUS-LIKE PARTICLE
Buffer solutionpH: 7.4
Buffer component

Buffer-ID: 1

IDConc.NameFormula
1218 mMsucrose
210 mMpotassium phosphate
325 mMcitrateCitric acid
4150 mMsodium chlorideNaClSodium chloride
525 mMHEPESC8H18N2O4S
61 mMsodium azideNaN3
SpecimenConc.: 0.33 mg/ml / Details: 0.33 mg/mL VLP + 15 mg/mL Mxra8 / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: 0.458 mbar.l/s O2 and 0.11 mbar.l/s H2 / Grid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R2/2
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277.15 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: OTHER / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy
Image recordingAverage exposure time: 0.3 sec. / Electron dose: 50 e/Å2 / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

EM software
IDNameVersionCategory
2RELION2.1image acquisition
4CTFFIND4CTF correction
7UCSF Chimera1.13.1model fitting
8Coot0.8.9.1model fitting
11FREALIGN10final Euler assignment
13cisTEM1.0.0-beta3D reconstruction
14PHENIX1.14model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: I (icosahedral)
3D reconstructionResolution: 4.06 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 22486 / Symmetry type: POINT
Atomic model buildingProtocol: FLEXIBLE FIT / Space: REAL
Atomic model building

3D fitting-ID: 1

IDPDB-IDPdb chain-IDPdb chain residue range
13N42B5-342
23N42F1-393
33J0CA394-442
43J0CB394-423
55H23A111-261
66NK3A39-291

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