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Basic information

Entry
Database: EMDB / ID: EMD-9366
TitleAtomic structures and deletion mutant reveal different capsid-binding patterns and functional significance of tegument protein pp150 in murine and human cytomegaloviruses with implications for therapeutic development
Map data3D reconstruction for MCMV capsid associated with pM32 tegument protein. Contour level=4.0 was estimated while showing the map using step=2 in Chimera.
Sample
  • Virus: Murid herpesvirus 1 (strain Smith)
    • Protein or peptide: Major capsid protein
    • Protein or peptide: Tegument protein
    • Protein or peptide: Small capsomere-interacting protein
    • Protein or peptide: Minor capsid protein
    • Protein or peptide: Triplex capsid protein 2
Function / homology
Function and homology information


T=16 icosahedral viral capsid / viral tegument / viral capsid assembly / viral process / viral capsid / host cell nucleus / structural molecule activity / DNA binding
Similarity search - Function
Herpesvirus UL11/UL32 / Small capsid protein, Herpesviridae / Herpesvirus large structural phosphoprotein UL32 / Small capsid protein of Herpesviridae / Herpesvirus capsid protein 2 / Herpesvirus capsid shell protein 1 / Herpesvirus VP23 like capsid protein / Herpesvirus capsid shell protein VP19C / Herpesvirus major capsid protein / Herpesvirus major capsid protein, upper domain superfamily / Herpes virus major capsid protein
Similarity search - Domain/homology
Major capsid protein / Tegument protein / Minor capsid protein / Small capsomere-interacting protein / Triplex capsid protein 2
Similarity search - Component
Biological speciesMuHV-1 (virus) / Murid herpesvirus 1 (strain Smith)
Methodsingle particle reconstruction / cryo EM / Resolution: 5.0 Å
AuthorsLiu W / Dai XH / Jih J / Chan K / Trang P / Yu XK / Balogun R / Mei Y / Liu FY / Zhou ZH
Funding support United States, 5 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Dental and Craniofacial Research (NIH/NIDCR)DE025567, DE023935, and DE025462 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM071940 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)AI069015 United States
National Science Foundation (NSF, United States)DMR-1548924 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)1S10RR23057 United States
CitationJournal: PLoS Pathog / Year: 2019
Title: Atomic structures and deletion mutant reveal different capsid-binding patterns and functional significance of tegument protein pp150 in murine and human cytomegaloviruses with implications for ...Title: Atomic structures and deletion mutant reveal different capsid-binding patterns and functional significance of tegument protein pp150 in murine and human cytomegaloviruses with implications for therapeutic development.
Authors: Wei Liu / Xinghong Dai / Jonathan Jih / Karen Chan / Phong Trang / Xuekui Yu / Rilwan Balogun / Ye Mei / Fenyong Liu / Z Hong Zhou /
Abstract: Cytomegalovirus (CMV) infection causes birth defects and life-threatening complications in immunosuppressed patients. Lack of vaccine and need for more effective drugs have driven widespread ongoing ...Cytomegalovirus (CMV) infection causes birth defects and life-threatening complications in immunosuppressed patients. Lack of vaccine and need for more effective drugs have driven widespread ongoing therapeutic development efforts against human CMV (HCMV), mostly using murine CMV (MCMV) as the model system for preclinical animal tests. The recent publication (Yu et al., 2017, DOI: 10.1126/science.aam6892) of an atomic model for HCMV capsid with associated tegument protein pp150 has infused impetus for rational design of novel vaccines and drugs, but the absence of high-resolution structural data on MCMV remains a significant knowledge gap in such development efforts. Here, by cryoEM with sub-particle reconstruction method, we have obtained the first atomic structure of MCMV capsid with associated pp150. Surprisingly, the capsid-binding patterns of pp150 differ between HCMV and MCMV despite their highly similar capsid structures. In MCMV, pp150 is absent on triplex Tc and exists as a "Λ"-shaped dimer on other triplexes, leading to only 260 groups of two pp150 subunits per capsid in contrast to 320 groups of three pp150 subunits each in a "Δ"-shaped fortifying configuration. Many more amino acids contribute to pp150-pp150 interactions in MCMV than in HCMV, making MCMV pp150 dimer inflexible thus incompatible to instigate triplex Tc-binding as observed in HCMV. While pp150 is essential in HCMV, our pp150-deletion mutant of MCMV remained viable though with attenuated infectivity and exhibiting defects in retaining viral genome. These results thus invalidate targeting pp150, but lend support to targeting capsid proteins, when using MCMV as a model for HCMV pathogenesis and therapeutic studies.
History
DepositionDec 22, 2018-
Header (metadata) releaseMar 6, 2019-
Map releaseMar 6, 2019-
UpdateDec 2, 2020-
Current statusDec 2, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 4
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 4
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6nhj
  • Surface level: 4
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-6nhj
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_9366.png

Downloads & links

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Map

FileDownload / File: emd_9366.map.gz / Format: CCP4 / Size: 7.8 GB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotation3D reconstruction for MCMV capsid associated with pM32 tegument protein. Contour level=4.0 was estimated while showing the map using step=2 in Chimera.
Voxel sizeX=Y=Z: 1.351 Å
Density
Contour LevelBy AUTHOR: 4 / Movie #1: 4
Minimum - Maximum-7.3179355 - 13.18449
Average (Standard dev.)0.00006514169 (±0.99999726)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-639-639-639
Dimensions128012801280
Spacing128012801280
CellA=B=C: 1729.2799 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.3511.3511.351
M x/y/z128012801280
origin x/y/z0.0000.0000.000
length x/y/z1729.2801729.2801729.280
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS-639-639-639
NC/NR/NS128012801280
D min/max/mean-7.31813.1840.000

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Supplemental data

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Additional map: Sub-particle reconstruction for MCMV 2-fold region

Fileemd_9366_additional_1.map
AnnotationSub-particle reconstruction for MCMV 2-fold region
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Sub-particle reconstruction for MCMV 3-fold region

Fileemd_9366_additional_2.map
AnnotationSub-particle reconstruction for MCMV 3-fold region
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Sub-particle reconstruction for MCMV 5-fold region

Fileemd_9366_additional_3.map
AnnotationSub-particle reconstruction for MCMV 5-fold region
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Murid herpesvirus 1 (strain Smith)

EntireName: Murid herpesvirus 1 (strain Smith)
Components
  • Virus: Murid herpesvirus 1 (strain Smith)
    • Protein or peptide: Major capsid protein
    • Protein or peptide: Tegument protein
    • Protein or peptide: Small capsomere-interacting protein
    • Protein or peptide: Minor capsid protein
    • Protein or peptide: Triplex capsid protein 2

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Supramolecule #1: Murid herpesvirus 1 (strain Smith)

SupramoleculeName: Murid herpesvirus 1 (strain Smith) / type: virus / ID: 1 / Parent: 0 / Macromolecule list: all / NCBI-ID: 10367 / Sci species name: Murid herpesvirus 1 (strain Smith) / Virus type: VIRION / Virus isolate: STRAIN / Virus enveloped: Yes / Virus empty: No

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Macromolecule #1: Major capsid protein

MacromoleculeName: Major capsid protein / type: protein_or_peptide / ID: 1 / Number of copies: 16 / Enantiomer: LEVO
Source (natural)Organism: MuHV-1 (virus) / Strain: Smith
Molecular weightTheoretical: 151.673031 KDa
SequenceString: MGENWTATEL LPKLDVPIDL LTHIKLSVGE EMFNNFRLYY GDDPERYNLS FEAIFGTYCN KIEWVTFLGT ALATAAHAIM FHDLNKMTT GKMLFYIQVP RVATGAGIPT SRQTTVMVSK YSEKSPITIP FEISAACLTH LKETFEDTLL DKLLNADAVN T VLRAVKNT ...String:
MGENWTATEL LPKLDVPIDL LTHIKLSVGE EMFNNFRLYY GDDPERYNLS FEAIFGTYCN KIEWVTFLGT ALATAAHAIM FHDLNKMTT GKMLFYIQVP RVATGAGIPT SRQTTVMVSK YSEKSPITIP FEISAACLTH LKETFEDTLL DKLLNADAVN T VLRAVKNT ADAMERGLID TFLRVLLRHA PPCFVLRTLM EHGTIARRMA TRVQRANIAQ GFKSKMLATI FLLDRSRDRG QL TRYLDML TDCVTESILD NPETYTVGGG ERLAGVIVST HTVVQALLNA LGGSIRRTGV KTPASYGKFV LSKENAVTAI AHH AIMADF SQHADRIQQS SQKDLPESQF LDQRLTFTET QMDVLKVGER LVALEHLRKV YKNTDVQDPL ERDVELTFYF PVGL HVPSG RAYSTAENKI KLVDTAENQL PTTVYFYNKD RIPQRISHAE ALKTLCHPAI HDAGPCLEAF AQAGPPQGDD RVRAL CRRE FVREHMAHAT RRLVHFYQAR IDPPRTANEA KHDFSTKEFA KVDNYLLFTE LHPFFDFCFH TENGQVRPLC TPRIMV GNL PEALAPADFH DLRAKQALEL TKVRAPEGHE ATLQVLRASL TDHQYPELFY LIESLIHGDP AAFETGIELV TRCVNNY WR QRGLLAFANS YDMVRLIATR LGDGAVVPAA YTHYRNLLSI TRFVARTCEL TGLNGRLCDE PLLAYVSALH DPRLWPPF V QALPRNANLV RVVADDVPLD AAHIEERNPG TSDVARMIAM DQAEPLFVDA RRTSDEEMVA QKVYYLCLVP AVLNNHACG AGLNLKHLLV KLFYTKFFLT ADPDSLTAGE EALTNNPLLA ALVRDVATDE NVTANQAAEE LFHLVAHVPE NAQMLEIRAA LDPAQRHGA PSAGFESLQH VLYNGFCMTT VPKLLQEYLT VIPFHRFYSD PGLAATANHD IRVFLNDFPQ YQRCDGGFPL S PIFAHEYH HWHRTPFSCY SAACAHTLES VLTLAIMHHK MSPVSIAALS RMGLHPGFAL TVVRTDTFET DTLLYSTKAS TA VIINTPI VTKEDRDINT VFHVSQNINT VEMGLGYGAT TCTAHLRRVR SDMGSRMQDL FQVFPMHVYR NEDVDAWVRQ ATG ARRTEV LDSEAISILT FGRKTDKGGP ALLHGQRATC EVILTPVSAD LEFFRYPNNP RGRSSSMLGV DPYDEDAALA TLYD HTSPD PQTFVSTNNP WGSQRGSLGD VIYNTRNREK LGHNPSFYSP CAQFFTTDDI INANKTLFKT VEEYLNRSQD CIHGE TDLQ YICVEGTNSI VEKPCRFLQE ALTQHTGTTQ ALMESQLKGT SKLGLDETHY GNYSIGETIP LQQSILFNS

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Macromolecule #2: Tegument protein

MacromoleculeName: Tegument protein / type: protein_or_peptide / ID: 2 / Number of copies: 8 / Enantiomer: LEVO
Source (natural)Organism: MuHV-1 (virus) / Strain: Smith
Molecular weightTheoretical: 78.749961 KDa
SequenceString: MSARGRAAGD DGRQAELMAT LGFVRLSKSS VGKVKKFLNN LYDLKSINLC RHPRVIAECR GTDLSRETQL YNEMVLWLRY HEKLTARRP GHLPLLTRIR QDYDKLFGFV AARPELCGFD GLTEVNVFDD AVYGDDYVPR VDVFLRGLED LARCLCAQGP D KPARAVIM ...String:
MSARGRAAGD DGRQAELMAT LGFVRLSKSS VGKVKKFLNN LYDLKSINLC RHPRVIAECR GTDLSRETQL YNEMVLWLRY HEKLTARRP GHLPLLTRIR QDYDKLFGFV AARPELCGFD GLTEVNVFDD AVYGDDYVPR VDVFLRGLED LARCLCAQGP D KPARAVIM GFINMRAEEV NRLMDNVRDA AERVLVYEVL DVRDPLNEDP SVLVHNRLVY LCRLAYAISK SWQTLSHMCL DR INSLRRR LILAFHDRPA FARVYARNAL ERPVDGTTAY NLLRRLEEDF LLFRNALRWG DPDWGLESEL ESEGDNSDAG SDL DLEDEE DDDDGGGPGG HDDESGGNRT PDPGMSLHDD TGIANTCLIG GDDEDCGGGS GRCLEFDPDS ERCLGVKMVN GRAL RWWNP TGIMVDNEAA VWIDEHGRVM DKPPPKELRK ASSDDGGNKK NPPPKKNVTP PVSGSNSVGG GVQTPSTASG KRTGK KKEG GGGYLLRSRS TDDDEVRKMK KDGTIDDRAD RELKMALQKA RESTADSDLS TILPRTEPLR KVAFVGDPVA AFGDTV RTT SSSKGFDDGP FTTQGASVLL PPPLGGPGST LTLPPDLPDL SGVLADEDVQ DSRYGKIPKS RTKKHPTFPE NYKQRPP PH NKDDEYYWDE TGDNMPVGED GGGVLEDLRK GLEGIDLKTG GGGSLQPPLS QQFAGSPFAG SDGDGGGLVK KSSSSH

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Macromolecule #3: Small capsomere-interacting protein

MacromoleculeName: Small capsomere-interacting protein / type: protein_or_peptide / ID: 3 / Number of copies: 16 / Enantiomer: LEVO
Source (natural)Organism: MuHV-1 (virus) / Strain: Smith
Molecular weightTheoretical: 9.845198 KDa
SequenceString:
MSTNVSSAAS GGGSSGGSSG ASSGGGGGGS GGSSKKEEER RKQFGANVLN LAPAMVAQPV ISTMIPKYMK MGGHEDKLAY QLDLLRMLS IAKKATVIQ

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Macromolecule #4: Minor capsid protein

MacromoleculeName: Minor capsid protein / type: protein_or_peptide / ID: 4 / Number of copies: 5 / Enantiomer: LEVO
Source (natural)Organism: MuHV-1 (virus) / Strain: Smith
Molecular weightTheoretical: 33.281289 KDa
SequenceString: MANVSSFGPM KREVMEFDPE DPYKVSKMRK LERSIAKGYV YGADHQAITA RFFVRESLGE VEQKNLGVLM FRLDTGIEMP STVLVSLFF LSMVAENVSA ATKNTLAAIY GREGEAIRTW LRDGAWRLHR VVHPLGCTNS ITPGATCLIT CSMRGHSYNM L KTEIYPLL ...String:
MANVSSFGPM KREVMEFDPE DPYKVSKMRK LERSIAKGYV YGADHQAITA RFFVRESLGE VEQKNLGVLM FRLDTGIEMP STVLVSLFF LSMVAENVSA ATKNTLAAIY GREGEAIRTW LRDGAWRLHR VVHPLGCTNS ITPGATCLIT CSMRGHSYNM L KTEIYPLL VPKEIYLDLD GESTDEIRFV YFVITYDYNS DRQGRPSAFV VVSRITHRHT LINVLRYRFR VSRFHFLNNS IS GYGPSTG CLGTLQRLGW FCSRDSRSGI VASRAGQLSV VKLEKFYVDV GPLVEFA

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Macromolecule #5: Triplex capsid protein 2

MacromoleculeName: Triplex capsid protein 2 / type: protein_or_peptide / ID: 5 / Number of copies: 10 / Enantiomer: LEVO
Source (natural)Organism: MuHV-1 (virus) / Strain: Smith
Molecular weightTheoretical: 34.660262 KDa
SequenceString: METTVLVTFE QRLTTGDVGK LSRLIGAVIP IPYRHHLLGS SQVGLDAVVK DKTRDYSRMR ARMREMTLTI MRRVEGNQMI LGVPTHGQC YTIRNTGPVS WEKGDVLTTL PPVFSGEVTG LVSVSDWDLV LPWIVPMALA TEINQRMMML ALLSLDRSHE E VRAATAQL ...String:
METTVLVTFE QRLTTGDVGK LSRLIGAVIP IPYRHHLLGS SQVGLDAVVK DKTRDYSRMR ARMREMTLTI MRRVEGNQMI LGVPTHGQC YTIRNTGPVS WEKGDVLTTL PPVFSGEVTG LVSVSDWDLV LPWIVPMALA TEINQRMMML ALLSLDRSHE E VRAATAQL RVVRYRDATL TLPEITIDDT VLIDMRNVCI SLSMIANLSS EVTLAYVRKL ALEDSNMLLM KCQEILGRRM PQ VGVGAGS SGDRNDPPAR SRTNYNITPT EELNKLTALF VMIRQITDVI SEQPAFLVCD VSPDDKSALC IYKG

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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: 7.4 / Details: PBS buffer, PH=7.4
GridMaterial: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 101.325 kPa
VitrificationCryogen name: ETHANE / Instrument: HOMEMADE PLUNGER / Details: The grids were manually plunged into the Ethane..

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated defocus max: 3.0 µm / Calibrated defocus min: 1.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 47000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: AGFA SCIENTA FILM / Digitization - Scanner: NIKON SUPER COOLSCAN 9000 / Number real images: 2200 / Average electron dose: 25.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 58254
CTF correctionSoftware - Name: CTFFIND
Startup modelType of model: OTHER / Details: a sphere density map
Initial angle assignmentType: COMMON LINE
Final angle assignmentType: COMMON LINE
Final reconstructionApplied symmetry - Point group: I (icosahedral) / Resolution.type: BY AUTHOR / Resolution: 5.0 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 47982

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: OTHER / Overall B value: 200
Output model

PDB-6nhj:
Atomic structures and deletion mutant reveal different capsid-binding patterns and functional significance of tegument protein pp150 in murine and human cytomegaloviruses with implications for therapeutic development

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