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- EMDB-11850: Atomic structure of the poxvirus initially transcribing complex i... -

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Basic information

Entry
Database: EMDB / ID: EMD-11850
TitleAtomic structure of the poxvirus initially transcribing complex in conformation 2
Map data
Sample
  • Complex: Vaccinia transcription pre-initiation complex
    • Complex: Vaccinia transcription pre-initiation complex
      • Protein or peptide: x 9 types
    • Complex: RNA
      • RNA: x 1 types
    • Complex: DNA
      • DNA: x 2 types
  • Ligand: x 2 types
KeywordsVaccinia / Virus / RNA polymerase / DNA-dependent / initiation complex / PIC / poxvirus / poxviridae / TRANSCRIPTION
Function / homology
Function and homology information


viral transcription / RNA polymerase I activity / DNA-directed RNA polymerase complex / nucleotidyltransferase activity / DNA-templated transcription termination / virion component / ribonucleoside binding / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / DNA-binding transcription factor activity ...viral transcription / RNA polymerase I activity / DNA-directed RNA polymerase complex / nucleotidyltransferase activity / DNA-templated transcription termination / virion component / ribonucleoside binding / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / DNA-binding transcription factor activity / DNA-templated transcription / DNA binding / zinc ion binding / metal ion binding
Similarity search - Function
RNA polymerase-associated transcription specificity factor Rap94 / RNA polymerase-associated transcription specificity factor, Rap94 / DNA-directed RNA polymerase, 18kDa subunit, poxviral / DNA-directed RNA polymerase, 35kDa subunit, poxviral / RNA polymerase, 22kDa subunit, poxviral / DNA-directed RNA polymerase, 19kDa subunit, poxviral / DNA-directed RNA polymerase, 7kDa polypeptide, chordopoxviral / RNA polymerase, 30kDa subunit, chordopox-type / RNA polymerase, 132kDa subunit, poxvirus-type / RNA polymerase, 30kDa subunit, chordopox-type, N-terminal ...RNA polymerase-associated transcription specificity factor Rap94 / RNA polymerase-associated transcription specificity factor, Rap94 / DNA-directed RNA polymerase, 18kDa subunit, poxviral / DNA-directed RNA polymerase, 35kDa subunit, poxviral / RNA polymerase, 22kDa subunit, poxviral / DNA-directed RNA polymerase, 19kDa subunit, poxviral / DNA-directed RNA polymerase, 7kDa polypeptide, chordopoxviral / RNA polymerase, 30kDa subunit, chordopox-type / RNA polymerase, 132kDa subunit, poxvirus-type / RNA polymerase, 30kDa subunit, chordopox-type, N-terminal / Poxvirus DNA-directed RNA polymerase, 18 kD subunit / Poxvirus DNA-directed RNA polymerase, 35 kD subunit / Poxvirus RNA polymerase 22 kDa subunit / Poxvirus DNA-directed RNA polymerase 19 kDa subunit / Chordopoxvirus DNA-directed RNA polymerase 7 kDa polypeptide (RPO7) / Poxvirus DNA dependent RNA polymerase 30kDa subunit / Poxvirus DNA dependent RNA polymerase / Zinc finger TFIIS-type signature. / RNA polymerase Rpb2, domain 5 / RNA polymerase Rpb2, domain 5 / Zinc finger, TFIIS-type / Transcription factor S-II (TFIIS) / Zinc finger TFIIS-type profile. / C2C2 Zinc finger / RPB6/omega subunit-like superfamily / RNA polymerase Rpb1, clamp domain superfamily / RNA polymerase Rpb1, domain 3 / RNA polymerase Rpb1, domain 3 / DNA-directed RNA polymerase, subunit beta-prime / RNA polymerase Rpb1, domain 1 / RNA polymerase Rpb1, domain 1 / RNA polymerase, alpha subunit / RNA polymerase Rpb1, domain 5 / RNA polymerase Rpb1, domain 4 / RNA polymerase Rpb1, domain 2 / RNA polymerase Rpb1, domain 5 / RNA polymerase Rpb1, domain 4 / RNA polymerase, N-terminal / RNA polymerase Rpb1, funnel domain superfamily / RNA polymerase I subunit A N-terminus / RNA polymerase, beta subunit, conserved site / RNA polymerase Rpb2, domain 7 / RNA polymerase Rpb2, domain 3 / RNA polymerase Rpb2, OB-fold / RNA polymerase Rpb2, domain 7 / RNA polymerase Rpb2, domain 3 / RNA polymerases beta chain signature. / DNA-directed RNA polymerase, subunit 2, hybrid-binding domain / DNA-directed RNA polymerase, subunit 2 / DNA-directed RNA polymerase, subunit 2, hybrid-binding domain superfamily / RNA polymerase Rpb2, domain 6
Similarity search - Domain/homology
DNA-directed RNA polymerase subunit / DNA-directed RNA polymerase 30 kDa polypeptide / RNA polymerase-associated transcription-specificity factor RAP94 / DNA-directed RNA polymerase 147 kDa polypeptide / DNA-directed RNA polymerase 35 kDa subunit / DNA-directed RNA polymerase / DNA-directed RNA polymerase 18 kDa subunit / DNA-directed RNA polymerase 19 kDa subunit / DNA-directed RNA polymerase 7 kDa subunit
Similarity search - Component
Biological speciesVaccinia virus GLV-1h68
Methodsingle particle reconstruction / cryo EM / Resolution: 3.15 Å
AuthorsGrimm C / Bartuli J
Funding support Germany, 1 items
OrganizationGrant numberCountry
German Research Foundation (DFG)Fi-573/20-1 Germany
CitationJournal: Nat Struct Mol Biol / Year: 2021
Title: Structural basis of the complete poxvirus transcription initiation process.
Authors: Clemens Grimm / Julia Bartuli / Bettina Boettcher / Aladar A Szalay / Utz Fischer /
Abstract: Poxviruses express their genes in the cytoplasm of infected cells using a virus-encoded multi-subunit polymerase (vRNAP) and unique transcription factors. We present cryo-EM structures that uncover ...Poxviruses express their genes in the cytoplasm of infected cells using a virus-encoded multi-subunit polymerase (vRNAP) and unique transcription factors. We present cryo-EM structures that uncover the complete transcription initiation phase of the poxvirus vaccinia. In the pre-initiation complex, the heterodimeric early transcription factor VETFs/l adopts an arc-like shape spanning the polymerase cleft and anchoring upstream and downstream promoter elements. VETFI emerges as a TBP-like protein that inserts asymmetrically into the DNA major groove, triggers DNA melting, ensures promoter recognition and enforces transcription directionality. The helicase VETFs fosters promoter melting and the phospho-peptide domain (PPD) of vRNAP subunit Rpo30 enables transcription initiation. An unprecedented upstream promoter scrunching mechanism assisted by the helicase NPH-I probably fosters promoter escape and transition into elongation. Our structures shed light on unique mechanisms of poxviral gene expression and aid the understanding of thus far unexplained universal principles in transcription.
History
DepositionOct 16, 2020-
Header (metadata) releaseOct 6, 2021-
Map releaseOct 6, 2021-
UpdateNov 20, 2024-
Current statusNov 20, 2024Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.026
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.026
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7ap8
  • Surface level: 0.026
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_11850.png

Downloads & links

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Map

FileDownload / File: emd_11850.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.06 Å/pix.
x 320 pix.
= 340.32 Å		1.06 Å/pix.
x 320 pix.
= 340.32 Å		1.06 Å/pix.
x 320 pix.
= 340.32 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.0635 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.026 / Movie #1: 0.026
Minimum - Maximum-0.035800166 - 0.11799699
Average (Standard dev.)0.000447742 (±0.004185777)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 340.32 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.06351.06351.0635
M x/y/z320320320
origin x/y/z0.0000.0000.000
length x/y/z340.320340.320340.320
α/β/γ90.00090.00090.000
start NX/NY/NZ-121-116-119
NX/NY/NZ240232241
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS320320320
D min/max/mean-0.0360.1180.000

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Supplemental data

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Sample components

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Entire : Vaccinia transcription pre-initiation complex

EntireName: Vaccinia transcription pre-initiation complex
Components
  • Complex: Vaccinia transcription pre-initiation complex
    • Complex: Vaccinia transcription pre-initiation complex
      • Protein or peptide: DNA-directed RNA polymerase 147 kDa polypeptide
      • Protein or peptide: DNA-directed RNA polymerase
      • Protein or peptide: DNA-directed RNA polymerase 35 kDa subunit
      • Protein or peptide: DNA-directed RNA polymerase subunit
      • Protein or peptide: DNA-directed RNA polymerase 19 kDa subunit
      • Protein or peptide: DNA-directed RNA polymerase 18 kDa subunit
      • Protein or peptide: Protein H4
      • Protein or peptide: DNA-directed RNA polymerase 7 kDa subunit
      • Protein or peptide: DNA-directed RNA polymerase 30 kDa polypeptide
    • Complex: RNA
      • RNA: RNA (5'-R(P*AP*UP*AP*AP*A)-3')
    • Complex: DNA
      • DNA: Synthetic promoter DNA oligomer, non-template strand
      • DNA: Synthetic promoter DNA oligomer, template strand
  • Ligand: MAGNESIUM ION
  • Ligand: ZINC ION

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Supramolecule #1: Vaccinia transcription pre-initiation complex

SupramoleculeName: Vaccinia transcription pre-initiation complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#12
Molecular weightTheoretical: 670 KDa

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Supramolecule #2: Vaccinia transcription pre-initiation complex

SupramoleculeName: Vaccinia transcription pre-initiation complex / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1-#8, #11
Source (natural)Organism: Vaccinia virus GLV-1h68

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Supramolecule #3: RNA

SupramoleculeName: RNA / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #10
Source (natural)Organism: Vaccinia virus GLV-1h68

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Supramolecule #4: DNA

SupramoleculeName: DNA / type: complex / ID: 4 / Parent: 1 / Macromolecule list: #9, #12
Source (natural)Organism: Vaccinia virus GLV-1h68

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Macromolecule #1: DNA-directed RNA polymerase 147 kDa polypeptide

MacromoleculeName: DNA-directed RNA polymerase 147 kDa polypeptide / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Vaccinia virus GLV-1h68
Molecular weightTheoretical: 146.995703 KDa
SequenceString: MAVISKVTYS LYDQKEINAT DIIISHVKND DDIGTVKDGR LGAMDGALCK TCGKTELECF GHWGKVSIYK THIVKPEFIS EIIRLLNHI CIHCGLLRSR EPYSDDINLK ELSGHALRRL KDKILSKKKS CWNSECMQPY QKITFSKKKV CFVNKLDDIN V PNSLIYQK ...String:
MAVISKVTYS LYDQKEINAT DIIISHVKND DDIGTVKDGR LGAMDGALCK TCGKTELECF GHWGKVSIYK THIVKPEFIS EIIRLLNHI CIHCGLLRSR EPYSDDINLK ELSGHALRRL KDKILSKKKS CWNSECMQPY QKITFSKKKV CFVNKLDDIN V PNSLIYQK LISIHEKFWP LLEIHQYPAN LFYTDYFPIP PLIIRPAISF WIDSIPKETN ELTYLLGMIV KNCNLNADEQ VI QKAVIEY DDIKIISNNT TSINLSYITS GKNNMIRSYI VARRKDQTAR SVIGPSTSIT VNEVGMPAYI RNTLTEKIFV NAF TVDKVK QLLASNQVKF YFNKRLNQLT RIRQGKFIKN KIHLLPGDWV EVAVQEYTSI IFGRQPSLHR YNVIASSIRA TEGD TIKIS PGIANSQNAD FDGDEEWMIL EQNPKAVIEQ SILMYPTTLL KHDIHGAPVY GSIQDEIVAA YSLFRIQDLC LDEVL NILG KYGREFDPKG KCKFSGKDIY TYLIGEKINY PGLLKDGEII ANDVDSNFVV AMRHLSLAGL LSDHKSNVEG INFIIK SSY VFKRYLSIYG FGVTFKDLRP NSTFTNKLEA INVEKIELIK EAYAKYLNDV RDGKIVPLSK ALEADYVESM LSNLTNL NI REIEEHMRQT LIDDPDNNLL KMAKAGYKVN PTELMYILGT YGQQRIDGEP AETRVLGRVL PYYLPDSKDP EGRGYILN S LTKGLTGSQY YFSMLVARSQ STDIVCETSR TGTLARKIIK KMEDMVVDGY GQVVIGNTLI KYAANYTKIL GSVCKPVDL IYPDESMTWY LEISALWNKI KQGFVYSQKQ KLAKKTLAPF NFLVFVKPTT EDNAIKVKDL YDMIHNVIDD VREKYFFTVS NIDFMEYIF LTHLNPSRIR ITKETAITIF EKFYEKLNYT LGGGTPIGII SAQVLSEKFT QQALSSFHTT EKSGAVKQKL G FNEFNNLT NLSKNKTEII TLVSDDISKL QSVKINFEFV CLGELNPNIT LRKETDKYVV DIIVNRLYIK RAEITELVVE YM IERFISF SVIVKEWGME TFIEDEDNIR FTVYLNFVEP EELNLSKFMM VLPGAANKGK ISKFKIPISD YTGYDDFNQT KKL NKMTVE LMNLKELGSF DLENVNVYPG VWNTYDIFGI EAAREYLCEA MLNTYGEGFD YLYQPCDLLA SLLCASYEPE SVNK FKFGA ASTLKRATFG DNKALLNAAL HKKSEPINDN SSCHFFSKVP NIGTGYYKYF IDLGLLMRME RKLSDKISSQ KIKEM EETE DF

UniProtKB: DNA-directed RNA polymerase 147 kDa polypeptide

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Macromolecule #2: DNA-directed RNA polymerase

MacromoleculeName: DNA-directed RNA polymerase / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Vaccinia virus GLV-1h68
Molecular weightTheoretical: 133.526859 KDa
SequenceString: MKKNTNSEMD QRLGYKFLVP DPKAGVFYRP LHFQYVSYSN FILHRLHEIL TVKRPLLSFK NNTERIMIEI SNVKVTPPDY SPIIASIKG KSYDALATFT VNIFKEVMTK EGISITKISS YEGKDSHLIK IPLLIGYGNK NPLDTAKYLV PNVIGGVFIN K QSVEKVGI ...String:
MKKNTNSEMD QRLGYKFLVP DPKAGVFYRP LHFQYVSYSN FILHRLHEIL TVKRPLLSFK NNTERIMIEI SNVKVTPPDY SPIIASIKG KSYDALATFT VNIFKEVMTK EGISITKISS YEGKDSHLIK IPLLIGYGNK NPLDTAKYLV PNVIGGVFIN K QSVEKVGI NLVEKITTWP KFRVVKPNSF TFSFSSVSPP NVLPTRYRHY KISLDISQLE ALNISSTKTF ITVNIVLLSQ YL SRVSLEF IRRSLSYDMP PEVVYLVNAI IDSAKRITES ITDFNIDTYI NDLVEAEHIK QKSQLTINEF KYEMLHNFLP HMN YTPDQL KGFYMISLLR KFLYCIFHTS RYPDRDSMVC HRILTYGKYF ETLAHDELEN YIGNIRNDIM NNHKNRGTYA VNIH VLTTP GLNHAFSSLL SGKFKKSDGS YRTHPHYSWM QNISIPRSVG FYPDQVKISK MFSVRKYHPS QYLYFCSSDV PERGP QVGL VSQLSVLSSI TNILTSEYLD LEKKICEYIR SYYKDDISYF ETGFPITIEN ALVASLNPNM ICDFVTDFRR RKRMGF FGN LEVGITLVRD HMNEIRINIG AGRLVRPFLV VDNGELMMDV CPELESRLDD MTFSDIQKEF PHVIEMVDIE QFTFSNV CE SVQKFRMMSK DERKQYDLCD FPAEFRDGYV ASSLVGINHN SGPRAILGCA QAKQAISCLS SDIRNKIDNG IHLMYPER P IVISKALETS KIAANCFGQH VTIALMSYKG INQEDGIIIK KQFIQRGGLD IVTAKKHQVE IPLENFNNKE RDRSNAYSK LESNGLVRLN AFLESGDAMA RNISSRTLED DFARDNQISF DVSEKYTDMY KSRVERVQVE LTDKVKVRVL TMKERRPILG DKFTTRTSQ KGTVAYVADE TELPYDENGI TPDVIINSTS IFSRKTISML IEVILTAAYS AKPYNNKGEN RPVCFPSSNE T SIDTYMQF AKQCYEHSNP KLSDEELSDK IFCEKILYDP ETDKPYASKV FFGPIYYLRL RHLTQDKATV RCRGKKTKLI RQ ANEGRKR GGGIKFGEME RDCLIAHGAA NTITEVLKDS EEDYQDVYVC ENCGDIAAQI KGINTCLRCS KLNLSPLLTK IDT THVSKV FLTQMNARGV KVKLDFERRP PSFYKPLDKV DLKPSFLV

UniProtKB: DNA-directed RNA polymerase

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Macromolecule #3: DNA-directed RNA polymerase 35 kDa subunit

MacromoleculeName: DNA-directed RNA polymerase 35 kDa subunit / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Vaccinia virus GLV-1h68
Molecular weightTheoretical: 35.430676 KDa
SequenceString: MQHPREENSI VVELEPSLAT FIKQGFNNLV KWPLLNIGIV LSNTSTAVNE EWLTAVEHIP TMKIFYKHIH KILTREMGFL VYLKRSQSE RDNYITLYDF DYYIIDKDTN SVTMVDKPTE LKETLLHVFQ EYRLKSSQTI ELIAFSSGTV INEDIVSKLT F LDVEVFNR ...String:
MQHPREENSI VVELEPSLAT FIKQGFNNLV KWPLLNIGIV LSNTSTAVNE EWLTAVEHIP TMKIFYKHIH KILTREMGFL VYLKRSQSE RDNYITLYDF DYYIIDKDTN SVTMVDKPTE LKETLLHVFQ EYRLKSSQTI ELIAFSSGTV INEDIVSKLT F LDVEVFNR EYNNVKTIID PDFVFRSPFI VISPMGKLTF FVEVYSWFDF KSCFKDIIDF LEGALIANIH NHMIKVGNCD ET VSSYNPE SGMLFVNDLM TMNIVNFFGC NSRLESYHRF DMTKVDVELF IKALSDACKK ILSASNRL

UniProtKB: DNA-directed RNA polymerase 35 kDa subunit

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Macromolecule #4: DNA-directed RNA polymerase subunit

MacromoleculeName: DNA-directed RNA polymerase subunit / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Vaccinia virus GLV-1h68
Molecular weightTheoretical: 21.36574 KDa
SequenceString:
MNQYNVKYLA KILCLKTEIA RDPYAVINRN VLLRYTTDIE YNDLVTLITV RHKIDSMKTV FQVFNESSIN YTPVDDDYGE PIIITSYLQ KGHNKFPVNF LYIDVVISDL FPSFVRLDTT ETNIVNSVLQ TGDGKKTLRL PKMLETEIVV KILYRPNIPL K IVRFFRNN MVTGVEIADR SVISVAD

UniProtKB: DNA-directed RNA polymerase subunit

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Macromolecule #5: DNA-directed RNA polymerase 19 kDa subunit

MacromoleculeName: DNA-directed RNA polymerase 19 kDa subunit / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Vaccinia virus GLV-1h68
Molecular weightTheoretical: 19.020088 KDa
SequenceString:
MADTDDIIDY ESDDLTEYED DEEEEEDGES LETSDIDPKS SYKIVESAST HIEDAHSNLK HIGNHISALK RRYTRRISLF EIAGIIAES YNLLQRGRLP LVSEFSDETM KQNMLHVIIQ EIEEGSCPIV IEKNGELLSV NDFDKDGLKF HLDYIIKIWK L QKRY

UniProtKB: DNA-directed RNA polymerase 19 kDa subunit

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Macromolecule #6: DNA-directed RNA polymerase 18 kDa subunit

MacromoleculeName: DNA-directed RNA polymerase 18 kDa subunit / type: protein_or_peptide / ID: 6 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Vaccinia virus GLV-1h68
Molecular weightTheoretical: 17.917195 KDa
SequenceString:
MSSFVTNGYL SVTLEPHELT LDIKTNIRNA VYKTYLHREI SGKMAKKIEI REDVELPLGE IVNNSVVINV PCVITYAYYH VGDIVRGTL NIEDESNVTI QCGDLICKLS RDSGTVSFSD SKYCFFRNGN AYDNGSEVTA VLMEAQQGIE SSFVFLANIV D S

UniProtKB: DNA-directed RNA polymerase 18 kDa subunit

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Macromolecule #7: Protein H4

MacromoleculeName: Protein H4 / type: protein_or_peptide / ID: 7 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Vaccinia virus GLV-1h68
Molecular weightTheoretical: 93.667633 KDa
SequenceString: MDSKETILIE IIPKIKAYLL DANISPKSYD DFISRNKNIF VINLYNVSTI TEEDIRLLYT TIEQNIDADD QTLVAIFSYI GYKFEQAVK EEISTSLSFN DKNTTDEMTY NLYDLFFNTL DMYLRQKKIS ILVNDDVRGD VIVSYKNSDL VSSFNAELEP E IKKIPFNM ...String:
MDSKETILIE IIPKIKAYLL DANISPKSYD DFISRNKNIF VINLYNVSTI TEEDIRLLYT TIEQNIDADD QTLVAIFSYI GYKFEQAVK EEISTSLSFN DKNTTDEMTY NLYDLFFNTL DMYLRQKKIS ILVNDDVRGD VIVSYKNSDL VSSFNAELEP E IKKIPFNM KNLLPYLEKN LDQLRFSKKY LDFAYLCRHI GIPISKKKYN VRYVFLYKID GLSIPIIIKD FLDVKYVYLE NT GKIYKNS FSEDHNNSLS DWGKVIIPLL KDRHLYSYIF LSSYHLHSYY TDLIARDEPV FVKRKKLDII EIDEPEAWKR DVR VEFAPC EHQIRLKEAM KVDANYFTKI NNFANEFIYY EDGVAYCRVC GINIPIFNLD AADVIKNTVI VSTFNKTIFL SEPY SYFVH SQRFIFNIIM SFDNIMKSQT WVMKYNINRL ILNFLIDINS RRQEYEKKFS SEIKRGLFFL RLSANLFESQ VSSTE LFYV SKMLNLNYIV ALVIILNSSA DFIVSYMTSK NKTVEESTLK YAISVVIYDF LVKTRICEKG SLDTIVLFTD VYTSIM PEE LDLHFQRITL ELRKLVSIQR SALEPNYDVE SRGEELPLSA LKFFDTSTII VKTMAPVHTC VEQKIVAPTP SVEPTDA SL KNFKELTCDE DIKILIRVHD TNATKLVIFP SHLKIEIERK KLIIPLKSLY ITNTLKYYYS NSYLYVFRFG DPMPFEEE L IDHEHVQYKI NCYNILRYHL LPDSDVFVYF SNSLNREALE YAFYIFLSKY VNVKQWIDEN ITRIKELYMI NFNN

UniProtKB: RNA polymerase-associated transcription-specificity factor RAP94

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Macromolecule #8: DNA-directed RNA polymerase 7 kDa subunit

MacromoleculeName: DNA-directed RNA polymerase 7 kDa subunit / type: protein_or_peptide / ID: 8 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Vaccinia virus GLV-1h68
Molecular weightTheoretical: 7.299715 KDa
SequenceString:
MVFQLVCSTC GKDISHERYK LIIRKKSLKD VLVSVKNECC RLKLSTQIEP QRNLTVQPLL DIN

UniProtKB: DNA-directed RNA polymerase 7 kDa subunit

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Macromolecule #11: DNA-directed RNA polymerase 30 kDa polypeptide

MacromoleculeName: DNA-directed RNA polymerase 30 kDa polypeptide / type: protein_or_peptide / ID: 11 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA-directed RNA polymerase
Source (natural)Organism: Vaccinia virus GLV-1h68
Molecular weightTheoretical: 29.834359 KDa
SequenceString: MENVYISSYS SNEQTSMAVA ATDIRELLSQ YVDDANLEDL IEWAMEKSSK YYIKNIGNTK SNIEETKFES KNNIGIEYSK DSRNKLSYR NKPSIATNLE YKTLCDMIKG TSGTEKEFLR YLLFGIKCIK KGVEYNIDKI KDVSYNDYFN VLDEKYNTPC P NCKSRNTT ...String:
MENVYISSYS SNEQTSMAVA ATDIRELLSQ YVDDANLEDL IEWAMEKSSK YYIKNIGNTK SNIEETKFES KNNIGIEYSK DSRNKLSYR NKPSIATNLE YKTLCDMIKG TSGTEKEFLR YLLFGIKCIK KGVEYNIDKI KDVSYNDYFN VLDEKYNTPC P NCKSRNTT PMMIQTRAAD EPPLVRHACR DCKQHFKPPK FRAFRNLNVT TQSIHENKEI TEILPDNNPS PPESPEPASP ID DGLIRAT FDRNDEPPED DE

UniProtKB: DNA-directed RNA polymerase 30 kDa polypeptide

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Macromolecule #9: Synthetic promoter DNA oligomer, non-template strand

MacromoleculeName: Synthetic promoter DNA oligomer, non-template strand / type: dna / ID: 9 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Vaccinia virus GLV-1h68
Molecular weightTheoretical: 18.417867 KDa
SequenceString: (DG)(DC)(DT)(DT)(DA)(DG)(DC)(DT)(DT)(DA) (DA)(DA)(DA)(DA)(DT)(DT)(DG)(DA)(DA)(DA) (DA)(DA)(DC)(DT)(DA)(DT)(DT)(DT)(DT) (DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DA) (DA)(DA)(DT)(DC)(DC)(DA) ...String:
(DG)(DC)(DT)(DT)(DA)(DG)(DC)(DT)(DT)(DA) (DA)(DA)(DA)(DA)(DT)(DT)(DG)(DA)(DA)(DA) (DA)(DA)(DC)(DT)(DA)(DT)(DT)(DT)(DT) (DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DA) (DA)(DA)(DT)(DC)(DC)(DA)(DT)(DA) (DA)(DC)(DC)(DC)(DA)(DC)(DT)(DT)(DG)(DG) (DG)(DG)

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Macromolecule #12: Synthetic promoter DNA oligomer, template strand

MacromoleculeName: Synthetic promoter DNA oligomer, template strand / type: dna / ID: 12 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Vaccinia virus GLV-1h68
Molecular weightTheoretical: 18.547941 KDa
SequenceString: (DC)(DC)(DC)(DC)(DA)(DA)(DG)(DT)(DG)(DG) (DG)(DT)(DT)(DA)(DT)(DG)(DG)(DA)(DT)(DT) (DT)(DA)(DT)(DA)(DG)(DA)(DA)(DT)(DT) (DT)(DA)(DG)(DA)(DA)(DA)(DT)(DA)(DG)(DT) (DT) (DT)(DT)(DT)(DC)(DA)(DA) ...String:
(DC)(DC)(DC)(DC)(DA)(DA)(DG)(DT)(DG)(DG) (DG)(DT)(DT)(DA)(DT)(DG)(DG)(DA)(DT)(DT) (DT)(DA)(DT)(DA)(DG)(DA)(DA)(DT)(DT) (DT)(DA)(DG)(DA)(DA)(DA)(DT)(DA)(DG)(DT) (DT) (DT)(DT)(DT)(DC)(DA)(DA)(DT)(DT) (DT)(DT)(DT)(DA)(DA)(DG)(DC)(DT)(DA)(DA) (DG)(DC)

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Macromolecule #10: RNA (5'-R(P*AP*UP*AP*AP*A)-3')

MacromoleculeName: RNA (5'-R(P*AP*UP*AP*AP*A)-3') / type: rna / ID: 10 / Number of copies: 1
Source (natural)Organism: Vaccinia virus GLV-1h68
Molecular weightTheoretical: 1.578032 KDa
SequenceString:
AUAAA

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Macromolecule #13: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 13 / Number of copies: 1 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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Macromolecule #14: ZINC ION

MacromoleculeName: ZINC ION / type: ligand / ID: 14 / Number of copies: 4 / Formula: ZN
Molecular weightTheoretical: 65.409 Da

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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

Concentration0.15 mg/mL
BufferpH: 7.5
GridModel: Quantifoil / Material: COPPER / Support film - Material: CARBON / Support film - topology: HOLEY
VitrificationCryogen name: ETHANE
Detailsmonodisperse sample prepared by sucrose gradient centrifugation

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 3 / Average exposure time: 74.99 sec. / Average electron dose: 78.9 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

6ric

Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.15 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 73035
Initial angle assignmentType: ANGULAR RECONSTITUTION
Final angle assignmentType: ANGULAR RECONSTITUTION

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