+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-43731 | |||||||||
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Title | Cryo-EM structure of BTV pre-core | |||||||||
Map data | ||||||||||
Sample |
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Keywords | Bluetongue virus / capsid protein / assembly intermediate / virion / VIRAL PROTEIN | |||||||||
Function / homology | Function and homology information viral genome replication / RNA-directed RNA polymerase / RNA-dependent RNA polymerase activity / nucleotide binding / DNA-templated transcription / structural molecule activity / RNA binding Similarity search - Function | |||||||||
Biological species | Bluetongue virus (serotype 1 / isolate South Africa) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.1 Å | |||||||||
Authors | Xia X / Sung PY / Martynowycz MW / Gonen T / Roy P / Zhou ZH | |||||||||
Funding support | United States, United Kingdom, 2 items
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Citation | Journal: Cell / Year: 2024 Title: RNA genome packaging and capsid assembly of bluetongue virus visualized in host cells. Authors: Xian Xia / Po-Yu Sung / Michael W Martynowycz / Tamir Gonen / Polly Roy / Z Hong Zhou / Abstract: Unlike those of double-stranded DNA (dsDNA), single-stranded DNA (ssDNA), and ssRNA viruses, the mechanism of genome packaging of dsRNA viruses is poorly understood. Here, we combined the techniques ...Unlike those of double-stranded DNA (dsDNA), single-stranded DNA (ssDNA), and ssRNA viruses, the mechanism of genome packaging of dsRNA viruses is poorly understood. Here, we combined the techniques of high-resolution cryoelectron microscopy (cryo-EM), cellular cryoelectron tomography (cryo-ET), and structure-guided mutagenesis to investigate genome packaging and capsid assembly of bluetongue virus (BTV), a member of the Reoviridae family of dsRNA viruses. A total of eleven assembly states of BTV capsid were captured, with resolutions up to 2.8 Å, with most visualized in the host cytoplasm. ATPase VP6 was found underneath the vertices of capsid shell protein VP3 as an RNA-harboring pentamer, facilitating RNA packaging. RNA packaging expands the VP3 shell, which then engages middle- and outer-layer proteins to generate infectious virions. These revealed "duality" characteristics of the BTV assembly mechanism reconcile previous contradictory co-assembly and core-filling models and provide insights into the mysterious RNA packaging and capsid assembly of Reoviridae members and beyond. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_43731.map.gz | 193.1 MB | EMDB map data format | |
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Header (meta data) | emd-43731-v30.xml emd-43731.xml | 20.1 KB 20.1 KB | Display Display | EMDB header |
Images | emd_43731.png | 113 KB | ||
Filedesc metadata | emd-43731.cif.gz | 7.3 KB | ||
Others | emd_43731_half_map_1.map.gz emd_43731_half_map_2.map.gz | 169.3 MB 169.3 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-43731 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-43731 | HTTPS FTP |
-Related structure data
Related structure data | 8w1sMC 8w12C 8w19C 8w1cC 8w1iC 8w1oC 8w1rC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_43731.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||
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Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.1 Å | ||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Half map: #2
File | emd_43731_half_map_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #1
File | emd_43731_half_map_2.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Sample components
-Entire : Bluetongue virus (serotype 1 / isolate South Africa)
Entire | Name: Bluetongue virus (serotype 1 / isolate South Africa) |
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Components |
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-Supramolecule #1: Bluetongue virus (serotype 1 / isolate South Africa)
Supramolecule | Name: Bluetongue virus (serotype 1 / isolate South Africa) / type: virus / ID: 1 / Parent: 0 / Macromolecule list: all / NCBI-ID: 10905 Sci species name: Bluetongue virus (serotype 1 / isolate South Africa) Virus type: VIRION / Virus isolate: SPECIES / Virus enveloped: No / Virus empty: No |
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Host (natural) | Organism: Ovis aries (sheep) |
-Macromolecule #1: Core protein VP3
Macromolecule | Name: Core protein VP3 / type: protein_or_peptide / ID: 1 / Number of copies: 10 / Enantiomer: LEVO |
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Source (natural) | Organism: Bluetongue virus (serotype 1 / isolate South Africa) |
Molecular weight | Theoretical: 103.410508 KDa |
Sequence | String: MAAQNEQRPE RIKTTPYLEG DVLSSDSGPL LSVFALQEIM QKVRQVQADY MTATREVDFT VPDVQKILDD IKALAAEQVY KIVKVPSIS FRHIVMQSRD RVLRVDTYYE EMSQVGDVIT EDEPEKFYST IIKKVRFIRG KGSFILHDIP TRDHRGMEVA E PEVLGVEF ...String: MAAQNEQRPE RIKTTPYLEG DVLSSDSGPL LSVFALQEIM QKVRQVQADY MTATREVDFT VPDVQKILDD IKALAAEQVY KIVKVPSIS FRHIVMQSRD RVLRVDTYYE EMSQVGDVIT EDEPEKFYST IIKKVRFIRG KGSFILHDIP TRDHRGMEVA E PEVLGVEF KNVLPVLTAE HRAMIQNALD GSIIENGNVA TRDVDVFIGA CSEPVYRIYN RLQGYIEAVQ LQELRNSIGW LE RLGHRKR ITYSQEVLTD FRRQDTIWVL ALQLPVNPQV VWDVPRSSIA NLIMNIATCL PTGEYIAPNP RISSITLTQR ITT TGPFAI LTGSTPTAQQ LNDVRKIYLA LMFPGQIILD LKIDPGERMD PAVRMVAGVV GHLLFTAGGR FTNLTQNMAR QLDI ALNDY LLYMYNTRVQ VNYGPTGEPL DFQIGRNQYD CNVFRADFAT GTGYNGWATI DVEYREPAPY VHAQRYIRYC GIDSR ELIN PTTYGIGMTY HCYNEMLRML VAAGKDSEAA YFRSMLPFHM VRFARINQII NEDLHSVFSL PDDMFNALLP DLIAGA HQN ADPVVLDVSW ISLWFAFNRS FEPTHRNEML EVAPLIESVY ASELSVMKVD MRHLSLMQRR FPDVLIQARP SHFWKAV LN DSPEAVKAVM NLSHSHNFIN IRDMMRWVML PSLQPSLKLA LEEEAWAAAN DFEDLMLTDQ VYMHRDMLPE PRLDDIER F RQEGFYYTNM LEAPPEIDRV VQYTYEIARL QANMGQFRAA LRRIMDDDDW VRFGGVLRTV RVKFYDARPP DDVLQGLPF SYDTNERGGL AYATIKYATE TTIFYLIYNV EFSNTPDSLV LINPTYTMTK VFINKRIVER VRVGQILAVL NRRFVAYKGK MRIMDITQS LKMGTKLAAP TV UniProtKB: Core protein VP3 |
-Macromolecule #2: RNA-directed RNA polymerase
Macromolecule | Name: RNA-directed RNA polymerase / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO / EC number: RNA-directed RNA polymerase |
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Source (natural) | Organism: Bluetongue virus (serotype 1 / isolate South Africa) |
Molecular weight | Theoretical: 149.926281 KDa |
Sequence | String: MVAITVQGAQ LIKRVVERFY PGIAFNINEG ACYIYKFSDH IRRIRMKHGT KYRRQAEEII RNISLRKERL YGIPVLDEVE WKYVFDGQT FQSYAFEVYV NSILPWSELD PEEEFLRNYR VSREMTEVEK FIEFRAKNEM QIYGDIPIKV WCCFINELSA E LKHVPLGM ...String: MVAITVQGAQ LIKRVVERFY PGIAFNINEG ACYIYKFSDH IRRIRMKHGT KYRRQAEEII RNISLRKERL YGIPVLDEVE WKYVFDGQT FQSYAFEVYV NSILPWSELD PEEEFLRNYR VSREMTEVEK FIEFRAKNEM QIYGDIPIKV WCCFINELSA E LKHVPLGM QVMADFVNRF DSPFHQGNRD LSNLEDFQVA YTTPLLFEMC CMESILEFNI KMRMREEEIS ALEFGDMKVD PV GLLREFF ILCLPHPKKI NNVLRAPYSW FVKMWGVGAD PIVVLQSTAG DDRNSKDVFY DKFRTEPNRY KALFRSSFYN ESR RMNEEK ILEAVKYSQK LGSHDRRLPL FEKMLKTVYT TPFYPHKSSN MILASFLLSI QTITGYGRAW VKNVSTEFDK QLKP NPSNL VQDVSDLTRE FFKQAYVEAK ERREEIVKPE DLYTSMLRLA RNTSSGFSTE IYVKKRFGPR LRDKDLIKIN SRIKA LVIF TKGHTVFTDE ELHKKYNSVE LYQTKGSRDV PIKATRTIYS INLSVLVPQL IVTLPLNEYF SRVGGITSPD YKKIGG KVI VGDLEATGSR VMDAADCFRN SADRDIFTIA IDYSEYDTHL TRHNFRTGML QGIREAMAPY RDLRYEGYTL EQIIDFG YG EGRVANTLWN GKRRLFKTTF DAYIRLDESE RDKGSFKVPK GVLPVSSVDV ANRIAVDKGF DTLIAATDGS DLALIDTH L SGENSTLIAN SMHNMAIGTL MQREVGREQP GVLTFLSEQY VGDDTLFYTK LHTTDTKVFD KVAASIFDTV AKCGHEASP SKTMMTPYSV EKTQTHAKQG CYVPQDRMMI ISSERRKDIE DVQGYVRSQV QTMITKVSRG FCHDLAQLIL MLKTTFIGAW KMKRTIKED AMYRDRKFDS NDEDGFTLIQ IRNPLALYVP IGWNGYGAHP AALNIVMTEE MYVDSIMISK LDEIMAPIRR I VHDIPPCW NETQGDKRGL ISATKMSFFS KMARPAVQAA LSDPQIINLV EELPLGEFSP GRISRTMMHS ALLKESSART LL SSGYELE YQKALNSWIT QVSMRLGEES GVISTSYAKL FDVYFEGELD GAPHMFPDQN LSPQFYIQKM MIGPRVSSRV RNS YVDRID VILRKDVVMR GFITANTILN VIEKLGTNHS VGDLVTVFTL MNIETRVAEE LAEYMTSEKI RFDALKLLKK GIAG DEFTM SLNVATQDFI DTYLAYPYQL TKTEVDAISL YCTQMIMLRA ALGLPKKKMK IVVTDDAKKR YKIRLQRFRT HVPKI KVLK KLIDPNRMTV RNLENQFV UniProtKB: RNA-directed RNA polymerase |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 8.8 / Component - Concentration: 20.0 mM / Component - Name: Tris |
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Grid | Model: PELCO Ultrathin Carbon with Lacey Carbon / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. / Pretreatment - Atmosphere: AIR |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 281.15 K / Instrument: FEI VITROBOT MARK IV |
Details | Virus isolated from infected BHK cells |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.6 µm / Nominal defocus min: 1.8 µm / Nominal magnification: 81000 |
Specialist optics | Energy filter - Name: GIF Quantum LS / Energy filter - Slit width: 20 eV |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 3 / Number real images: 62928 / Average exposure time: 2.0 sec. / Average electron dose: 50.0 e/Å2 |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
Particle selection | Number selected: 559848 |
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Startup model | Type of model: OTHER |
Initial angle assignment | Type: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1) |
Final 3D classification | Number classes: 7 / Avg.num./class: 500000 / Software - Name: RELION (ver. 3.1) |
Final angle assignment | Type: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1) |
Final reconstruction | Number classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 324633 |
-Atomic model buiding 1
Refinement | Space: REAL / Protocol: AB INITIO MODEL / Overall B value: 98 |
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Output model | PDB-8w1s: |