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- EMDB-7954: Fako virus full particles, projection-matching reconstruction wit... -

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Basic information

Entry
Database: EMDB / ID: EMD-7954
TitleFako virus full particles, projection-matching reconstruction without imposed symmetry
Map dataUnsymmetrized reconstruction of Fako virus full particles
Sample
  • Virus: Fako virus
    • Protein or peptide: clamp protein
    • Protein or peptide: major capsid protein
    • Protein or peptide: turret protein
    • Protein or peptide: RNA-dependent RNA polymerase
    • Protein or peptide: polymerase accessory protein
Biological speciesFako virus
Methodsingle particle reconstruction / cryo EM / Resolution: 10.7 Å
AuthorsKaelber JT / Jiang W / Weaver SC / Auguste AJ / Chiu W
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical SciencesP41GM103832 United States
Welch FoundationQ1242 United States
CitationJournal: Structure / Year: 2020
Title: Arrangement of the Polymerase Complexes inside a Nine-Segmented dsRNA Virus.
Authors: Jason T Kaelber / Wen Jiang / Scott C Weaver / Albert J Auguste / Wah Chiu /
Abstract: Members of the family Reoviridae package several copies of the viral polymerase complex into their capsid to carry out replication and transcription within viral particles. Classical single-particle ...Members of the family Reoviridae package several copies of the viral polymerase complex into their capsid to carry out replication and transcription within viral particles. Classical single-particle reconstruction encounters difficulties resolving structures such as the intraparticle polymerase complex because refinement can converge to an incorrect map and because the map could depict a nonrepresentative subset of particles or an average of heterogeneous particles. Using the nine-segmented Fako virus, we tested hypotheses for the arrangement and number of polymerase complexes within the virion by measuring how well each hypothesis describes the set of cryoelectron microscopy images of individual viral particles. We find that the polymerase complex in Fako virus binds at ten possible sites despite having only nine genome segments. A single asymmetric configuration describes the arrangement of these complexes in both virions and genome-free capsids. Similarities between the arrangements of Reoviridae with 9, 10, and 11 segments indicate the generalizability of this architecture.
History
DepositionMay 29, 2018-
Header (metadata) releaseJan 16, 2019-
Map releaseJan 16, 2019-
UpdateAug 5, 2020-
Current statusAug 5, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 2.62
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 2.62
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_7954.png

Downloads & links

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Map

FileDownload / File: emd_7954.map.gz / Format: CCP4 / Size: 729 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationUnsymmetrized reconstruction of Fako virus full particles
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.51 Å/pix.
x 576 pix.
= 867.456 Å		1.51 Å/pix.
x 576 pix.
= 867.456 Å		1.51 Å/pix.
x 576 pix.
= 867.456 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.506 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 2.62 / Movie #1: 2.62
Minimum - Maximum-6.0184875 - 7.844518
Average (Standard dev.)0.047352515 (±0.757196)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-288-288-288
Dimensions576576576
Spacing576576576
CellA=B=C: 867.45605 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.5061.5061.506
M x/y/z576576576
origin x/y/z0.0000.0000.000
length x/y/z867.456867.456867.456
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ352352352
MAP C/R/S123
start NC/NR/NS-288-288-288
NC/NR/NS576576576
D min/max/mean-6.0187.8450.047

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Supplemental data

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Half map: Unsymmetrized reconstruction of Fako virus full particles. Unmasked,...

Fileemd_7954_half_map_1.map
AnnotationUnsymmetrized reconstruction of Fako virus full particles. Unmasked, unfiltered half-set map.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Unsymmetrized reconstruction of Fako virus full particles. Unmasked,...

Fileemd_7954_half_map_2.map
AnnotationUnsymmetrized reconstruction of Fako virus full particles. Unmasked, unfiltered half-set map.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Fako virus

EntireName: Fako virus
Components
  • Virus: Fako virus
    • Protein or peptide: clamp protein
    • Protein or peptide: major capsid protein
    • Protein or peptide: turret protein
    • Protein or peptide: RNA-dependent RNA polymerase
    • Protein or peptide: polymerase accessory protein

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Supramolecule #1: Fako virus

SupramoleculeName: Fako virus / type: virus / ID: 1 / Parent: 0 / Macromolecule list: all / NCBI-ID: 1567404 / Sci species name: Fako virus / Sci species strain: CSW77 / Virus type: VIRION / Virus isolate: STRAIN / Virus enveloped: No / Virus empty: No
Host (natural)Organism: Culicinae (mosquito)
Molecular weightTheoretical: 43 MDa
Virus shellShell ID: 1 / T number (triangulation number): 2

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Macromolecule #1: clamp protein

MacromoleculeName: clamp protein / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Fako virus / Strain: CSW77
SequenceString: MTLTYWDKEK RMTLKQMIQQ VAINEQENEL THYVFTTPLS MPTFGKPMLG YVPLNEVATS KFFSNVNDFD RDNQLAMAH FPDTTITQAY NLTNSIKPGD TSLPDAEVAA LKWFWKFFTS INLVRQPPMD NVMYWACQFL S SGTSFLPL ERDVEIVFSG FKGSHICMFS ...String:
MTLTYWDKEK RMTLKQMIQQ VAINEQENEL THYVFTTPLS MPTFGKPMLG YVPLNEVATS KFFSNVNDFD RDNQLAMAH FPDTTITQAY NLTNSIKPGD TSLPDAEVAA LKWFWKFFTS INLVRQPPMD NVMYWACQFL S SGTSFLPL ERDVEIVFSG FKGSHICMFS NLRQMNLSPI LCPYYDLITN FKTTTEIRAY VDAHEELKSL LT YLCLCTI VGLCDTFTET RNMDTGEYVW KVRDVVSRNH TPAQNVEKFC YTIQNAKYMI QLVHVLLFPL TDN KYADLP NYVAVITQGA INQSRSHNVI NTTDESNSNT TSDTAASTSG IVSGDTGTVA SLYPDEFKYV QS

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Macromolecule #2: major capsid protein

MacromoleculeName: major capsid protein / type: protein_or_peptide / ID: 2 / Enantiomer: LEVO
Source (natural)Organism: Fako virus / Strain: CSW77
SequenceString: MRPIRMYKNN QERTNLKHQE INEEQQNEQT TSNQGFTRSD NSGKINIERI SSSRNQITDG KTVSSYSKIE TNRSSQDSV QHGGSSITYT SDTTGNPRIT NARTNNDETH ATGPIEDLNS TSHGREPEIE SFADRAELAM M IQGMTVGA LTVQPMRSIR STFANLANVL ...String:
MRPIRMYKNN QERTNLKHQE INEEQQNEQT TSNQGFTRSD NSGKINIERI SSSRNQITDG KTVSSYSKIE TNRSSQDSV QHGGSSITYT SDTTGNPRIT NARTNNDETH ATGPIEDLNS TSHGREPEIE SFADRAELAM M IQGMTVGA LTVQPMRSIR STFANLANVL IFHDVFTTED KPSAFIEYHS DEMIVNMPKQ TYNPIDNLAK IL YLPSLEK FKYGTGIVQL NYSPHISKLY QNTNNIINTI TDGITYANRT EFFIRVMVLM MMDRKILTME FYD VDTSAI SNTAILPTIP TTTGVSPLLR IDTRTEPIWY NDAIKTLITN LTIQYGKIKT VLDANAVKRY SVVG YPIDQ YRAYLYNHNL LEYLGKKVKR EDIMSLIKAL SYEFDLITIS DLEYQNIPKW FSDNDLSRFI FSICM FPDI VRQFHALNID YFSQANVFTV KSENAIVKML NSNQNMEPTI INWFLFRICA IDKTVIDDYF SLEMTP IIM RPKLYDFDMK RGEPVSLLYI LELILFSIMF PNVTQHMLGQ IQARILYISM YAFRQEYLKF ITKFGFY YK IVNGRKEYIQ VTNQNERMTE NNDVLTGNLY PSLFTDDPTL SAIAPTLAKI ARLMKPTTSL TPDDRAIA A KFPRFKDSAH LNPYSSLNIG GRTQHSVTYT RMYDAIEEMF NLILRAFASS FAQRPRAGVT QLKSLLTQL ADPLCLALDG HVYHLYNVMA NMMQNFIPNT DGQFHSFRAC SYAVKDGGNI YRVVQNGDEL NESLLIDTAI VWGLLGNTD SSYGNAIGAT GTANVPTKVQ PVIPTPDNFI TPTIHLKTSI DAICSVEGIL LLILSRQTTI P GYEDELNK LRTGISQPKV TERQYRRARE SIKNMLGSGD YNVAPLHFLL HTEHRSTKLS KPLIRRVLDN VV QPYVANL DPAEFENTPQ LIENSNMTRL QIALKMLTGD MDDIVKGLIL HKRACAKFDV YETLTIPTDV KTI VLTMQH ISTQTQNNMV YYVFLIDGVK ILAEDIKNVN FQIDITGIWP EYVITLLLRA INNGFNTYVS MPNI LYKPT ITADVRQFMN TTKAETLLIS NKSIVHEIMF FDNALQPKMS SDTLALSEAV YRTIWNSSII TQRIS ARGL MNLEDARPPE AKISHQSELD MGKIDETSGE PIYTSGLQKM QSSKVSMANV VLSAGSDVIR QAAIKY NVV RTQEIILFE

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Macromolecule #3: turret protein

MacromoleculeName: turret protein / type: protein_or_peptide / ID: 3 / Enantiomer: LEVO
Source (natural)Organism: Fako virus / Strain: CSW77
SequenceString: MIDLRLEEDI LTATLPEFLS TRPKYRYAYT NTKQQDIRFQ GPMRHVRLTH LYKQTKLWNL QYIERELAIS EIDDALDEF IQTFSLPYVI EQGTYKYNML LGMHAHNVNY QDDVSELIAN NPQLLNYLDD NPFSAIFELV N VDLQIYQY GQNIFNNEAE HTILFLKDNT ...String:
MIDLRLEEDI LTATLPEFLS TRPKYRYAYT NTKQQDIRFQ GPMRHVRLTH LYKQTKLWNL QYIERELAIS EIDDALDEF IQTFSLPYVI EQGTYKYNML LGMHAHNVNY QDDVSELIAN NPQLLNYLDD NPFSAIFELV N VDLQIYQY GQNIFNNEAE HTILFLKDNT NYGVIQALQK HPFSATHINW HLHKHIFVFH SREQLLNKLL SA GLEDSQL YQRQKTYSTK RGDRPTERMV TYIEDDHIRR IQAVFPLLLD NIFDVKLHKD SSMTWLKSYA DMI YDSVKN SNSTITPEIR KLYLRMYNQY MRIFLPIEQY MLYDNTCWPF SEKITLKINV RLISSRENQP VLWK TPIDT ENLISIVQPD EPINKLNFTA IPSTMIRLND NITMYRAVKD MFSAIEYLPD AIENIPTLTM KEQAL SRYI SPDSEAQNFF NNQPPYLNSI MNVNRQVFEA VKRGNIQVST GSMEHLCLCM HVKSGLIVGR TVLIDD KVV LRRNFNASTA KMITCYVKAF AQLYGEGSLI NPGLRMVFFG VETEPAIDIL KLFYGDKSLY IQGFGDR GI GRDKFRTKIE DALTLRIGCD ILISDIDQAD YEDPNEEKFD DITDFVCYVT ELVISNATVG LVKISMPT Y YIMNKISSTL NNKFSNVAIN IVKLSTQKPY TYEAYIMLSH GSTLTNKGYL RNPVCDVYLE KISLQPMDL KIISTISNEI NYDKPTLYRF VVDKNDVTDV SIAMHILSIH CSTITTRSVM VRSDNTGAFV TMSGIKDMKR VAIMNRMTD GTSANSYMHE QNGKLYLQKV PYLEDLISAF PNGFGSTYQN DYDSSMSVIN VNALIRQVVY R VISKSIPV ALLESLSRIR IIGGRDLGEM NAVYKLYKTP IEVYDAVGIT REYPHVQISY RAQRYSFTES IP NHTLLLA NYVIMNDVDG APISSLEQIN TIKKIISKIS LGSIAYIQVY TDIVARNINV MTKNDSFLIS ANA DKTVFK VQVSGYKAVE MCNYEQLLQL VSDNTGVNII KLTYQDVLES CVLSSGILGD TGSWLLDLVL ASTY IIEIR G

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Macromolecule #4: RNA-dependent RNA polymerase

MacromoleculeName: RNA-dependent RNA polymerase / type: protein_or_peptide / ID: 4 / Enantiomer: LEVO
Source (natural)Organism: Fako virus / Strain: CSW77
SequenceString: MATEPNEINL RMTQTLMKQI EDTTFKIRRK HDDMYTYLID DTAAIEMYPD TETPFRMKSS LHELYIQVKC QIPKSIIRP PLYNNDMYPL NNNNTNFLED RPFNYLCNFD WYEFLSRTKD ELGMHYNMIR DLIAMSSQTR Y SNIWSNIS GIIMYLRQYQ RGFALRAILA ...String:
MATEPNEINL RMTQTLMKQI EDTTFKIRRK HDDMYTYLID DTAAIEMYPD TETPFRMKSS LHELYIQVKC QIPKSIIRP PLYNNDMYPL NNNNTNFLED RPFNYLCNFD WYEFLSRTKD ELGMHYNMIR DLIAMSSQTR Y SNIWSNIS GIIMYLRQYQ RGFALRAILA RLLKQWNNFP FFDTWDGFRD VLPNTSTAWP LLFYAFMSVT FD YICENIS EGEAIVCFQN HLENAQVSYQ DTKLEKKNNY TLWLKYECHN LRVALTPRYD YTGTVSGNFL KDT TEYIIE NEFENNEIAM RNNLLPSFYA ILQQMRQKLK TVEDIIRLLT ICYSARDDRT YYGTLMELAI SKAI KPQVA GSIVPAPIPT SWLQKDPKMV LSARYPSTSF LSQMQEFYRR YYPALQKEID VHALSASFIN FLSTA SAGV GIELPEEIVN MVQDKRLLYL IKKGSGKRVL QEALLVEKYN DLDPIINDFV RLIKIVVRKQ IERRQR GIA GIPNNVLKIN QVTYEANKPF SKIARAPSHG KQSGNASDIH DLLFYTTQED VSHIETNGKR QMRGIVI SS ADVKGMDTHI QINAAMNQHL GAIEILDGIQ YDVGPFRQTN AIIQDIQGNV YEKSLNGGQQ AIAFGLAN F SQTTGINSKY FGQIPNQEGT FPSGLITTSN HHTQMLTLLI ETALTTFTKE FGKSMAISHL MILGDDVSL MLHGNDKDIN FFMKYLVEKF SQLGLILERD ESRNFGVFLQ QHVINGRFNG FSNRIAIFTS EDYKTRKSVR ESCTEYNAL IDDVIFRTYN VRKLLQFQRI HQFVVLSKYV FRIQNYKYES LRAKLAKRMN VFEYELKIKD N DKTDVHNR NALRFIGIQI PYTYFQYSGG GEIPPESFQK KDGSFTYEYS IYSPRGKWLR KFLYDISHDA RD VKFQIDH EMMKLYNLDI CDFLLQYNVL DIQEEIRSTI IDRELVSKLA MNLESLEHSN ARMISRRASE SLR SMGIRL PANGVYGYQI NERLVKVLQN IQQSDYEVKM VGDTLFTAIM EKFEHHKVRM EKGDKLHNFT LDFS NKDDK ITINRKMLAL HNISISKNMM PYSDAWMLYT CLDNTYNTSS DLATALAHSQ GHFKSFQYDR DMFAE SVKI ASKHGIGSLP MELFFEASNI RDTAQLKWIE AIKYYVQFKD YLYPYSLNPR RLFFIPEQVS SVSNIL NQE NIPADINQKA LLFRRAYAYV LSHPFCISGA KAIFVEDRI

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Macromolecule #5: polymerase accessory protein

MacromoleculeName: polymerase accessory protein / type: protein_or_peptide / ID: 5 / Enantiomer: LEVO
Source (natural)Organism: Fako virus / Strain: CSW77
SequenceString: MFAIPFTASQ VYSHLGLYQQ PIEKLDPLTK VQYESAEKLR DIITQLSRRS GGISLRKNLI RDIAFGNKPK LNIPSLTHQ VRHNREFKHG RGQFILVDSN GSDYYDGIFD DMVLILRSNF VLPNVPGIIS EQLRLIVIDN D FYLKHIPS EIAILRDVLS EHMVKVSIDG ...String:
MFAIPFTASQ VYSHLGLYQQ PIEKLDPLTK VQYESAEKLR DIITQLSRRS GGISLRKNLI RDIAFGNKPK LNIPSLTHQ VRHNREFKHG RGQFILVDSN GSDYYDGIFD DMVLILRSNF VLPNVPGIIS EQLRLIVIDN D FYLKHIPS EIAILRDVLS EHMVKVSIDG VSLDDPYISK FFRKPFILYD HTKFRKVDLT KVVLINKTGK SK AQLIDKY KLTDDAVFIS YEVFDIMLQP VKSRDGDIEQ FVRMRGDIET WKMKIGTTFE SNLMELFIHG VPV MNSTSQ LQLDFQISDI RSANVFDEQI VCAAKLNGLR EQCLKESSLI RVNIIGQKGS GKSMLMRLIN ERGI PGISR KIICIDSDAY GKWKTQTQYL DDKFSALKLI NVDNLHEISQ DDNIISYYEQ FIIDQLLAHN ITISE HTNS IRFIKQFKVD TLKDIGRKFK ELYLADFKEQ IHFYEYLCGN IPEPSSTLLI TFLHATVETS AAPGTN MNF SLNTILYPLQ SILNRKRGAL VNFILNRIYD EMGTEAFTRL RPCDVWK

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 7.8
Component:
ConcentrationFormulaName
10.0 mMC4H12NO3ClTris-HCl
1.0 mMEDTA
100.0 mMNaClsodium chloride
VitrificationCryogen name: ETHANE / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeJEOL 3200FSC
Specialist opticsEnergy filter - Name: In-column Omega Filter
Image recordingFilm or detector model: DIRECT ELECTRON DE-20 (5k x 3k) / Detector mode: INTEGRATING / Number real images: 2400 / Average electron dose: 30.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 4.7 mm
Sample stageSpecimen holder model: JEOL 3200FSC CRYOHOLDER / Cooling holder cryogen: NITROGEN

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Image processing

Particle selectionNumber selected: 10588
CTF correctionSoftware - Name: jspr
Startup modelType of model: EMDB MAP
EMDB ID:

6001

Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 10.7 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: jspr / Number images used: 5294
Initial angle assignmentType: OTHER / Software - Name: jspr
Final angle assignmentType: PROJECTION MATCHING / Software - Name: jspr
FSC plot (resolution estimation)

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