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- EMDB-34927: Cryo-EM structure of monkeypox virus DNA replication holoenzyme F... -

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Basic information

Entry
Database: EMDB / ID: EMD-34927
TitleCryo-EM structure of monkeypox virus DNA replication holoenzyme F8, A22 and E4 complex without DNA at 2.76 angostram
Map data
Sample
  • Complex: Monkeypox virus replication holoenzyme F8-A22-E4 complex without DNA
    • Protein or peptide: DNA polymerase
    • Protein or peptide: DNA polymerase processivity factor component A20
    • Protein or peptide: E4R
KeywordsDNA replication machinery / DNA polymerase / B-family DNA polymerase / uracil-DNA glycosylase / MPXV / orthopoxvirus / poxviridae / DNA processivity factor / VIRAL PROTEIN
Function / homology
Function and homology information


viral DNA genome replication / uracil DNA N-glycosylase activity / DNA recombination / DNA replication / DNA-directed DNA polymerase / DNA-directed DNA polymerase activity / nucleotide binding / DNA repair / DNA binding
Similarity search - Function
DNA-directed DNA polymerase, family B, viral insert domain / DNA polymerase B exonuclease, N-terminal / DNA polymerase family B viral insert / DNA polymerase family B exonuclease domain, N-terminal / Chordopoxvirus A20R / Chordopoxvirus A20R protein / Uracil-DNA glycosylase, active site / Uracil-DNA glycosylase signature. / Uracil-DNA glycosylase-like domain superfamily / DNA polymerase family B signature. ...DNA-directed DNA polymerase, family B, viral insert domain / DNA polymerase B exonuclease, N-terminal / DNA polymerase family B viral insert / DNA polymerase family B exonuclease domain, N-terminal / Chordopoxvirus A20R / Chordopoxvirus A20R protein / Uracil-DNA glycosylase, active site / Uracil-DNA glycosylase signature. / Uracil-DNA glycosylase-like domain superfamily / DNA polymerase family B signature. / DNA-directed DNA polymerase, family B, conserved site / DNA polymerase family B / DNA polymerase family B, exonuclease domain / DNA-directed DNA polymerase, family B, exonuclease domain / DNA-directed DNA polymerase, family B, multifunctional domain / DNA polymerase, palm domain superfamily / DNA polymerase type-B family / DNA-directed DNA polymerase, family B / Ribonuclease H superfamily / Ribonuclease H-like superfamily / DNA/RNA polymerase superfamily
Similarity search - Domain/homology
DNA polymerase processivity factor / Uracil-DNA glycosylase / DNA polymerase
Similarity search - Component
Biological speciesMonkeypox virus
Methodsingle particle reconstruction / cryo EM / Resolution: 2.76 Å
AuthorsXu Y / Wu Y / Zhang Y / Fan R / Yang Y / Li D / Yang B / Zhang Z / Dong C / Zhang X ...Xu Y / Wu Y / Zhang Y / Fan R / Yang Y / Li D / Yang B / Zhang Z / Dong C / Zhang X / Tang X / Dong H
Funding support China, 1 items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)32250710142 China
CitationJournal: To Be Published
Title: Structure of DNA replication machinery from human monkeypox virus
Authors: Xu YX / Wu YQ / Zhang YY / Fan RX / Yang YX / Li DY / Yang B / Zhang ZY / Dong CJ
History
DepositionDec 11, 2022-
Header (metadata) releaseDec 13, 2023-
Map releaseDec 13, 2023-
UpdateDec 13, 2023-
Current statusDec 13, 2023Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_34927.png

Downloads & links

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Map

FileDownload / File: emd_34927.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 0.84 Å
Density
Contour LevelBy AUTHOR: 0.06
Minimum - Maximum-1.1994206 - 1.751165
Average (Standard dev.)-0.000307297 (±0.023979208)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 268.8 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_34927_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_34927_half_map_2.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 : Monkeypox virus replication holoenzyme F8-A22-E4 complex without DNA

EntireName: Monkeypox virus replication holoenzyme F8-A22-E4 complex without DNA
Components
  • Complex: Monkeypox virus replication holoenzyme F8-A22-E4 complex without DNA
    • Protein or peptide: DNA polymerase
    • Protein or peptide: DNA polymerase processivity factor component A20
    • Protein or peptide: E4R

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Supramolecule #1: Monkeypox virus replication holoenzyme F8-A22-E4 complex without DNA

SupramoleculeName: Monkeypox virus replication holoenzyme F8-A22-E4 complex without DNA
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all / Details: F8-A22-E4 complex
Source (natural)Organism: Monkeypox virus
Molecular weightTheoretical: 191 kDa/nm

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Macromolecule #1: DNA polymerase

MacromoleculeName: DNA polymerase / type: protein_or_peptide / ID: 1
Details: MPXV DNA replication machinery catalytic subunit F8
Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Monkeypox virus
Molecular weightTheoretical: 120.041156 KDa
Recombinant expressionOrganism: Insect expression vector pBlueBacmsGCA1His (others)
SequenceString: MHHHHHHHHD YDIPTTENLY FQGMDVRCIN WFESHGENRF LYLKSRCRNG ETVFIRFPHY FYYVVTDEIY QSLSPPPFNA RPMGKMRTI DIDETISYNL DIKDRKCSVA DMWLIEEPKK RSIQNATMDE FFNISWFYIS NGISPDGCYS LDEQYLTKIN N GCYHCDDP ...String:
MHHHHHHHHD YDIPTTENLY FQGMDVRCIN WFESHGENRF LYLKSRCRNG ETVFIRFPHY FYYVVTDEIY QSLSPPPFNA RPMGKMRTI DIDETISYNL DIKDRKCSVA DMWLIEEPKK RSIQNATMDE FFNISWFYIS NGISPDGCYS LDEQYLTKIN N GCYHCDDP RNCFAKEIPR FDIPRSYLFL DIECHFDKKF PSVFINPISH TSYCYIDLSG KRLLFTLINE EMLTEQEIQE AV DRGCLRI QSLMEMDYER ELVLCSEIVL LRIAKQLLEL TFDYVVTFNG HNFDLRYITN RLELLTGEKI IFRSPDKKEA VHL CIYERN QSSHKGVCGM ANTTFHVNNN NGTIFFDLYS FIQKSEKLDS YKLDSISKNA FSCMGKVLNR GVREMTFIGD DTTD AKGKA DTFAKVLTTG NYVTVDEDII CKVIRKDILE NGFKVVLSCP TLPNDIYKLS FGKDDIDLAQ MYKDYNLNIA LDMAR YCIH DACLCQYLWE YYGVETKTDA GAATYVLPQS MVFEYRASTI IKGPLLKLLL ETKTILVRSE TKQKFPYEGG KVFAPK QKM FSNNVLIFDY NSLYPNVCIF GNLSPETLVG VVVSTNRLEE EINNQLLLQK YPPPRYITVH CEPRLPNLIS EIAIFDR SI EGTIPRLLRT FLAERARYKK MLKQATSSTE KAIYDSMQYT YKIVANSVYG LMGFRNSALY SYASAKSCTS IGRRMILY L ESVLNGAELS NGMLRFANTL SNPFYMDDRD INPIVKTSLP IDYRFRFRSV YGDTDSVFTE IDSQDVDKSI EIAKELERL INSRVLFNNF KIEFEAVYKN LIMQSKKKYT TMKYSASSNS KSVPERINKG TSETRRDVSK FHKNMIKTYK TRLSEMLSEG RMNSNQVCI DILRSLETDL RSEFDSRSSP LELFMLSRMH HSNYKSADNP NMYLVTEYNK NNPETIELGE RYYFAYICPA N VPWTKKLV NIKTYETIID RSFKLGSNQR IFYEVYFKRL TSEIVNLLDN KVLCISFFQR MFGSRPTFYE A

UniProtKB: DNA polymerase

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Macromolecule #2: DNA polymerase processivity factor component A20

MacromoleculeName: DNA polymerase processivity factor component A20 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Monkeypox virus
Molecular weightTheoretical: 49.203926 KDa
Recombinant expressionOrganism: Insect expression vector pBlueBachsGCA1 (others)
SequenceString: MTSSADLTNL KELLSLYKSL RFSDSVAIEK YNSLVEWGTS TYWKIGVQKV TNVETSISDY YDEVKNKPFN IDPGYYIFLP VYFGSVFIY SKGKNMVELG SGNSFQIPDE IRSACNKVLD SDNGIDFLRF VLLNNRWIME DAISKYQSPV NIFKLASEYG L NIPNYLEI ...String:
MTSSADLTNL KELLSLYKSL RFSDSVAIEK YNSLVEWGTS TYWKIGVQKV TNVETSISDY YDEVKNKPFN IDPGYYIFLP VYFGSVFIY SKGKNMVELG SGNSFQIPDE IRSACNKVLD SDNGIDFLRF VLLNNRWIME DAISKYQSPV NIFKLASEYG L NIPNYLEI EIEEDTLFDD ELYSIMERSF DDTFPKISIS YIKLGELKRQ VVDFFKFSFM YIESIKVDRI GDNIFIPSVI TK SGKKILV KDVDHLIRSK VREHTFVKVK KKNTFSILYD YDGNGTETRG EVIKRIIDTI GRDYYVNGKY FSKVGIAGLK QLT NKLDIN ECATVDELVD EINKSGTVKR KIKNQSVFDL SRECLGYPEA DFITLVNNMR FKIENCKVVN FNIENTNCLN NPSI ETIYG NFNQFVSIFN TVTDVKKRLF E

UniProtKB: DNA polymerase processivity factor

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Macromolecule #3: E4R

MacromoleculeName: E4R / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO / EC number: uracil-DNA glycosylase
Source (natural)Organism: Monkeypox virus
Molecular weightTheoretical: 27.883709 KDa
Recombinant expressionOrganism: Insect expression vector pBlueBachsGCA1 (others)
SequenceString: MHHHHHHDYD IPTTENLYFQ GASMNSVTIS HAPYTITYHD DWEPVMSQLV EFYNEVASWL LRDETSPIPD KFFIQLKQPL RNKRVCVCG IDPYPKDGTG VPFESPNFTK KSIKEIASSI SRLTGVIDYK GYNLNIIDGV IPWNYYLSCK LGETKSHAIY W DKISKLLL ...String:
MHHHHHHDYD IPTTENLYFQ GASMNSVTIS HAPYTITYHD DWEPVMSQLV EFYNEVASWL LRDETSPIPD KFFIQLKQPL RNKRVCVCG IDPYPKDGTG VPFESPNFTK KSIKEIASSI SRLTGVIDYK GYNLNIIDGV IPWNYYLSCK LGETKSHAIY W DKISKLLL QHITKHVSVL YCLGKTDFSN IRAKLESPVT TIVGYHPAAR DHQFEKDRSF EIINVLLELD NKTPINWAQG FI Y

UniProtKB: Uracil-DNA glycosylase

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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: 8
Details: 25mM HEPES pH 8.0, 300mM NaCl, 5% (w/v) glycerol, and 1 mM TCEP
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE
DetailsThis is a complex of F8-A22-E4

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Electron microscopy

MicroscopeTFS KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 3.0 µm / Nominal defocus min: 1.0 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 50.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionDetails: 11,662,000
Startup modelType of model: OTHER / Details: Alphfold2 prediction
Initial angle assignmentType: NOT APPLICABLE
Final 3D classificationNumber classes: 6 / Software - Name: cryoSPARC
Final angle assignmentType: NOT APPLICABLE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.76 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 343570
DetailsThe selected movies are corrected with motion correct

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