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- EMDB-37345: Yeast replisome in state IV -

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Basic information

Entry
Database: EMDB / ID: EMD-37345
TitleYeast replisome in state IV
Map data
Sample
  • Complex: replisome complex in state IV
    • Protein or peptide: x 13 types
    • DNA: x 1 types
Keywordsreplisome / complex / DNA replication / REPLICATION
Function / homology
Function and homology information


establishment of sister chromatid cohesion / DNA-templated DNA replication maintenance of fidelity / Unwinding of DNA / DNA replication initiation / Cul8-RING ubiquitin ligase complex / epsilon DNA polymerase complex / DNA strand elongation involved in mitotic DNA replication / MCM core complex / Assembly of the pre-replicative complex / Switching of origins to a post-replicative state ...establishment of sister chromatid cohesion / DNA-templated DNA replication maintenance of fidelity / Unwinding of DNA / DNA replication initiation / Cul8-RING ubiquitin ligase complex / epsilon DNA polymerase complex / DNA strand elongation involved in mitotic DNA replication / MCM core complex / Assembly of the pre-replicative complex / Switching of origins to a post-replicative state / nuclear DNA replication / MCM complex binding / GINS complex / mitotic DNA replication preinitiation complex assembly / premeiotic DNA replication / pre-replicative complex assembly involved in nuclear cell cycle DNA replication / mitotic DNA replication / Activation of the pre-replicative complex / CMG complex / nuclear pre-replicative complex / Activation of ATR in response to replication stress / MCM complex / DNA replication preinitiation complex / double-strand break repair via break-induced replication / replication fork protection complex / mitotic DNA replication initiation / single-stranded DNA helicase activity / regulation of DNA-templated DNA replication initiation / silent mating-type cassette heterochromatin formation / DNA strand elongation involved in DNA replication / mitotic sister chromatid cohesion / nuclear chromosome / DNA unwinding involved in DNA replication / DNA replication origin binding / nuclear replication fork / subtelomeric heterochromatin formation / DNA replication initiation / error-prone translesion synthesis / heterochromatin formation / DNA helicase activity / helicase activity / DNA-templated DNA replication / nucleosome assembly / single-stranded DNA binding / mitotic cell cycle / DNA helicase / chromosome, telomeric region / hydrolase activity / cell cycle / DNA repair / chromatin binding / ATP hydrolysis activity / DNA binding / nucleoplasm / ATP binding / identical protein binding / metal ion binding / nucleus / cytoplasm
Similarity search - Function
DNA polymerase epsilon, subunit B / : / DNA polymerase alpha-binding protein Ctf4, C-terminal domain / Minichromosome loss protein Mcl1, middle region / Minichromosome loss protein, Mcl1, middle region / CDC45 family / CDC45-like protein / DNA replication complex GINS protein Psf2 / GINS complex, subunit Psf1 / GINS complex, subunit Psf3 ...DNA polymerase epsilon, subunit B / : / DNA polymerase alpha-binding protein Ctf4, C-terminal domain / Minichromosome loss protein Mcl1, middle region / Minichromosome loss protein, Mcl1, middle region / CDC45 family / CDC45-like protein / DNA replication complex GINS protein Psf2 / GINS complex, subunit Psf1 / GINS complex, subunit Psf3 / GINS complex, subunit Psf3 superfamily / DNA replication complex GINS protein SLD5, C-terminal / GINS, helical bundle-like domain superfamily / GINS complex protein Sld5, alpha-helical domain / DNA replication complex GINS protein SLD5 C-terminus / GINS complex subunit Sld5 / GINS subunit, domain A / GINS complex protein / MCM4, winged helix domain / DNA polymerase alpha/delta/epsilon, subunit B / DNA polymerase alpha/epsilon subunit B / DNA replication licensing factor Mcm5 / DNA replication licensing factor Mcm3 / Mini-chromosome maintenance complex protein 4 / DNA replication licensing factor Mcm6 / DNA replication licensing factor Mcm7 / Mcm6, C-terminal winged-helix domain / MCM6 C-terminal winged-helix domain / DNA replication licensing factor Mcm2 / Mini-chromosome maintenance protein 2 / Mini-chromosome maintenance, conserved site / MCM family signature. / MCM N-terminal domain / MCM N-terminal domain / MCM OB domain / MCM OB domain / Mini-chromosome maintenance protein / MCM, AAA-lid domain / MCM P-loop domain / MCM AAA-lid domain / MCM family domain profile. / minichromosome maintenance proteins / MCM domain / WD40-repeat-containing domain superfamily / WD40/YVTN repeat-like-containing domain superfamily / Winged helix-like DNA-binding domain superfamily / ATPases associated with a variety of cellular activities / AAA+ ATPase domain / Nucleic acid-binding, OB-fold / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
DNA replication licensing factor MCM2 / DNA replication licensing factor MCM3 / DNA polymerase epsilon subunit B / Minichromosome maintenance protein 5 / DNA replication licensing factor MCM4 / DNA replication licensing factor MCM7 / DNA replication complex GINS protein PSF2 / DNA replication licensing factor MCM6 / DNA polymerase alpha-binding protein / DNA replication complex GINS protein SLD5 ...DNA replication licensing factor MCM2 / DNA replication licensing factor MCM3 / DNA polymerase epsilon subunit B / Minichromosome maintenance protein 5 / DNA replication licensing factor MCM4 / DNA replication licensing factor MCM7 / DNA replication complex GINS protein PSF2 / DNA replication licensing factor MCM6 / DNA polymerase alpha-binding protein / DNA replication complex GINS protein SLD5 / Cell division control protein 45 / DNA replication complex GINS protein PSF3 / DNA replication complex GINS protein PSF1
Similarity search - Component
Biological speciesSaccharomyces cerevisiae (brewer's yeast) / Saccharomyces cerevisiae S288C (yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 7.39 Å
AuthorsDang S / Zhai Y / Feng J / Yu D
Funding support Hong Kong, 8 items
OrganizationGrant numberCountry
The University Grants Committee, Research Grants Council (RGC)GRF16104617 Hong Kong
The University Grants Committee, Research Grants Council (RGC)GRF16103918 Hong Kong
The University Grants Committee, Research Grants Council (RGC)GRF17112119 Hong Kong
The University Grants Committee, Research Grants Council (RGC)GRF17101720 Hong Kong
The University Grants Committee, Research Grants Council (RGC)C7028-19GF Hong Kong
The University Grants Committee, Research Grants Council (RGC)C7009-20GF Hong Kong
The University Grants Committee, Research Grants Council (RGC)ECS26101919 Hong Kong
The University Grants Committee, Research Grants Council (RGC)GRF16103321 Hong Kong
CitationJournal: Nat Commun / Year: 2023
Title: Synergism between CMG helicase and leading strand DNA polymerase at replication fork.
Authors: Zhichun Xu / Jianrong Feng / Daqi Yu / Yunjing Huo / Xiaohui Ma / Wai Hei Lam / Zheng Liu / Xiang David Li / Toyotaka Ishibashi / Shangyu Dang / Yuanliang Zhai /
Abstract: The replisome that replicates the eukaryotic genome consists of at least three engines: the Cdc45-MCM-GINS (CMG) helicase that separates duplex DNA at the replication fork and two DNA polymerases, ...The replisome that replicates the eukaryotic genome consists of at least three engines: the Cdc45-MCM-GINS (CMG) helicase that separates duplex DNA at the replication fork and two DNA polymerases, one on each strand, that replicate the unwound DNA. Here, we determined a series of cryo-electron microscopy structures of a yeast replisome comprising CMG, leading-strand polymerase Polε and three accessory factors on a forked DNA. In these structures, Polε engages or disengages with the motor domains of the CMG by occupying two alternative positions, which closely correlate with the rotational movement of the single-stranded DNA around the MCM pore. During this process, the polymerase remains stably coupled to the helicase using Psf1 as a hinge. This synergism is modulated by a concerted rearrangement of ATPase sites to drive DNA translocation. The Polε-MCM coupling is not only required for CMG formation to initiate DNA replication but also facilitates the leading-strand DNA synthesis mediated by Polε. Our study elucidates a mechanism intrinsic to the replisome that coordinates the activities of CMG and Polε to negotiate any roadblocks, DNA damage, and epigenetic marks encountered during translocation along replication forks.
History
DepositionAug 31, 2023-
Header (metadata) releaseDec 20, 2023-
Map releaseDec 20, 2023-
UpdateDec 20, 2023-
Current statusDec 20, 2023Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_37345.png

Downloads & links

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Map

FileDownload / File: emd_37345.map.gz / Format: CCP4 / Size: 325 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.06 Å
Density
Contour LevelBy AUTHOR: 0.45
Minimum - Maximum-0.45040664 - 1.4159375
Average (Standard dev.)0.009585416 (±0.06352371)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions440440440
Spacing440440440
CellA=B=C: 466.39996 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_37345_msk_1.map
Projections & Slices
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Half map: #2

Fileemd_37345_half_map_1.map
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Half map: #1

Fileemd_37345_half_map_2.map
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Sample components

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Entire : replisome complex in state IV

EntireName: replisome complex in state IV
Components
  • Complex: replisome complex in state IV
    • Protein or peptide: DNA replication licensing factor MCM2
    • Protein or peptide: DNA replication licensing factor MCM3
    • Protein or peptide: DNA replication licensing factor MCM4
    • Protein or peptide: Minichromosome maintenance protein 5
    • Protein or peptide: DNA replication licensing factor MCM6
    • Protein or peptide: DNA replication licensing factor MCM7
    • Protein or peptide: DNA replication complex GINS protein PSF1
    • Protein or peptide: DNA replication complex GINS protein PSF2
    • Protein or peptide: DNA replication complex GINS protein PSF3
    • Protein or peptide: DNA replication complex GINS protein SLD5
    • Protein or peptide: Cell division control protein 45
    • Protein or peptide: DNA polymerase alpha-binding protein
    • DNA: DNA (71-mer)
    • Protein or peptide: DNA polymerase epsilon subunit B

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Supramolecule #1: replisome complex in state IV

SupramoleculeName: replisome complex in state IV / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#14
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Molecular weightTheoretical: 1.70 MDa

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Macromolecule #1: DNA replication licensing factor MCM2

MacromoleculeName: DNA replication licensing factor MCM2 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Molecular weightTheoretical: 98.911539 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)
SequenceString: MSDNRRRRRE EDDSDSENEL PPSSPQQHFR GGMNPVSSPI GSPDMINPEG DDNEVDDVPD IDEVEEQMNE VDLMDDNMYE DYAADHNRD RYDPDQVDDR EQQELSLSER RRIDAQLNER DRLLRNVAYI DDEDEEQEGA AQLDEMGLPV QRRRRRRQYE D LENSDDDL ...String:
MSDNRRRRRE EDDSDSENEL PPSSPQQHFR GGMNPVSSPI GSPDMINPEG DDNEVDDVPD IDEVEEQMNE VDLMDDNMYE DYAADHNRD RYDPDQVDDR EQQELSLSER RRIDAQLNER DRLLRNVAYI DDEDEEQEGA AQLDEMGLPV QRRRRRRQYE D LENSDDDL LSDMDIDPLR EELTLESLSN VKANSYSEWI TQPNVSRTIA RELKSFLLEY TDETGRSVYG ARIRTLGEMN SE SLEVNYR HLAESKAILA LFLAKCPEEM LKIFDLVAME ATELHYPDYA RIHSEIHVRI SDFPTIYSLR ELRESNLSSL VRV TGVVTR RTGVFPQLKY VKFNCLKCGS ILGPFFQDSN EEIRISFCTN CKSKGPFRVN GEKTVYRNYQ RVTLQEAPGT VPPG RLPRH REVILLADLV DVSKPGEEVE VTGIYKNNYD GNLNAKNGFP VFATIIEANS IKRREGNTAN EGEEGLDVFS WTEEE EREF RKISRDRGII DKIISSMAPS IYGHRDIKTA VACSLFGGVP KNVNGKHSIR GDINVLLLGD PGTAKSQILK YVEKTA HRA VFATGQGASA VGLTASVRKD PITKEWTLEG GALVLADKGV CLIDEFDKMN DQDRTSIHEA MEQQSISISK AGIVTTL QA RCSIIAAANP NGGRYNSTLP LAQNVSLTEP ILSRFDILCV VRDLVDEEAD ERLATFVVDS HVRSHPENDE DREGEELK N NGESAIEQGE DEINEQLNAR QRRLQRQRKK EEEISPIPQE LLMKYIHYAR TKIYPKLHQM DMDKVSRVYA DLRRESIST GSFPITVRHL ESILRIAESF AKMRLSEFVS SYDLDRAIKV VVDSFVDAQK VSVRRQLRRS FAIYTLGH

UniProtKB: DNA replication licensing factor MCM2

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Macromolecule #2: DNA replication licensing factor MCM3

MacromoleculeName: DNA replication licensing factor MCM3 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA helicase
Source (natural)Organism: Saccharomyces cerevisiae S288C (yeast) / Strain: ATCC 204508 / S288c
Molecular weightTheoretical: 107.653508 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)
SequenceString: MEGSTGFDGD ATTFFAPDAV FGDRVRRFQE FLDTFTSYRD SVRSIQVYNS NNAANYNDDQ DDADERDLLG DDDGDDLEKE KKAASSTSL NILPHRIIIS LDDLREFDRS FWSGILVEPA YFIPPAEKAL TDLADSMDDV PHPNASAVSS RHPWKLSFKG S FGAHALSP ...String:
MEGSTGFDGD ATTFFAPDAV FGDRVRRFQE FLDTFTSYRD SVRSIQVYNS NNAANYNDDQ DDADERDLLG DDDGDDLEKE KKAASSTSL NILPHRIIIS LDDLREFDRS FWSGILVEPA YFIPPAEKAL TDLADSMDDV PHPNASAVSS RHPWKLSFKG S FGAHALSP RTLTAQHLNK LVSVEGIVTK TSLVRPKLIR SVHYAAKTGR FHYRDYTDAT TTLTTRIPTP AIYPTEDTEG NK LTTEYGY STFIDHQRIT VQEMPEMAPA GQLPRSIDVI LDDDLVDKTK PGDRVNVVGV FKSLGAGGMN QSNSNTLIGF KTL ILGNTV YPLHARSTGV AARQMLTDFD IRNINKLSKK KDIFDILSQS LAPSIYGHDH IKKAILLMLM GGVEKNLENG SHLR GDINI LMVGDPSTAK SQLLRFVLNT ASLAIATTGR GSSGVGLTAA VTTDRETGER RLEAGAMVLA DRGVVCIDEF DKMTD VDRV AIHEVMEQQT VTIAKAGIHT TLNARCSVIA AANPVFGQYD VNRDPHQNIA LPDSLLSRFD LLFVVTDDIN EIRDRS ISE HVLRTHRYLP PGYLEGEPVR ERLNLSLAVG EDADINPEEH SNSGAGVENE GEDDEDHVFE KFNPLLQAGA KLAKNKG NY NGTEIPKLVT IPFLRKYVQY AKERVIPQLT QEAINVIVKN YTDLRNDDNT KKSPITARTL ETLIRLATAH AKVRLSKT V NKVDAKVAAN LLRFALLGED IGNDIDEEES EYEEALSKRS PQKSPKKRQR VRQPASNSGS PIKSTPRRST ASSVNATPS SARRILRFQD DEQNAGEDDN DIMSPLPADE EAELQRRLQL GLRVSPRRRE HLHAPEEGSS GPLTEVGTPR LPNVSSAGQD DEQQQSVIS FDNVEPGTIS TGRLSLISGI IARLMQTEIF EEESYPVASL FERINEELPE EEKFSAQEYL AGLKIMSDRN N LMVADDKV WRV

UniProtKB: DNA replication licensing factor MCM3

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Macromolecule #3: DNA replication licensing factor MCM4

MacromoleculeName: DNA replication licensing factor MCM4 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA helicase
Source (natural)Organism: Saccharomyces cerevisiae S288C (yeast) / Strain: ATCC 204508 / S288c
Molecular weightTheoretical: 105.138375 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)
SequenceString: MSQQSSSPTK EDNNSSSPVV PNPDSVPPQL SSPALFYSSS SSQGDIYGRN NSQNLSQGEG NIRAAIGSSP LNFPSSSQRQ NSDVFQSQG RQGRIRSSAS ASGRSRYHSD LRSDRALPTS SSSLGRNGQN RVHMRRNDIH TSDLSSPRRI VDFDTRSGVN T LDTSSSSA ...String:
MSQQSSSPTK EDNNSSSPVV PNPDSVPPQL SSPALFYSSS SSQGDIYGRN NSQNLSQGEG NIRAAIGSSP LNFPSSSQRQ NSDVFQSQG RQGRIRSSAS ASGRSRYHSD LRSDRALPTS SSSLGRNGQN RVHMRRNDIH TSDLSSPRRI VDFDTRSGVN T LDTSSSSA PPSEASEPLR IIWGTNVSIQ ECTTNFRNFL MSFKYKFRKI LDEREEFINN TTDEELYYIK QLNEMRELGT SN LNLDARN LLAYKQTEDL YHQLLNYPQE VISIMDQTIK DCMVSLIVDN NLDYDLDEIE TKFYKVRPYN VGSCKGMREL NPN DIDKLI NLKGLVLRST PVIPDMKVAF FKCNVCDHTM AVEIDRGVIQ EPARCERIDC NEPNSMSLIH NRCSFADKQV IKLQ ETPDF VPDGQTPHSI SLCVYDELVD SCRAGDRIEV TGTFRSIPIR ANSRQRVLKS LYKTYVDVVH VKKVSDKRLD VDTST IEQE LMQNKVDHNE VEEVRQITDQ DLAKIREVAA REDLYSLLAR SIAPSIYELE DVKKGILLQL FGGTNKTFTK GGRYRG DIN ILLCGDPSTS KSQILQYVHK ITPRGVYTSG KGSSAVGLTA YITRDVDTKQ LVLESGALVL SDGGVCCIDE FDKMSDS TR SVLHEVMEQQ TISIAKAGII TTLNARSSIL ASANPIGSRY NPNLPVTENI DLPPPLLSRF DLVYLVLDKV DEKNDREL A KHLTNLYLED KPEHISQDDV LPVEFLTMYI SYAKEHIHPI ITEAAKTELV RAYVGMRKMG DDSRSDEKRI TATTRQLES MIRLAEAHAK MKLKNVVELE DVQEAVRLIR SAIKDYATDP KTGKIDMNLV QTGKSVIQRK LQEDLSREIM NVLKDQASDS MSFNELIKQ INEHSQDRVE SSDIQEALSR LQQEDKVIVL GEGVRRSVRL NNRV

UniProtKB: DNA replication licensing factor MCM4

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Macromolecule #4: Minichromosome maintenance protein 5

MacromoleculeName: Minichromosome maintenance protein 5 / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA helicase
Source (natural)Organism: Saccharomyces cerevisiae S288C (yeast) / Strain: ATCC 204508 / S288c
Molecular weightTheoretical: 86.505734 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)
SequenceString: MSFDRPEIYS APVLQGESPN DDDNTEIIKS FKNFILEFRL DSQFIYRDQL RNNILVKNYS LTVNMEHLIG YNEDIYKKLS DEPSDIIPL FETAITQVAK RISILSRAQS ANNNDKDPEN TSMDTDSLLL NSLPTFQLIL NSNANQIPLR DLDSEHVSKI V RLSGIIIS ...String:
MSFDRPEIYS APVLQGESPN DDDNTEIIKS FKNFILEFRL DSQFIYRDQL RNNILVKNYS LTVNMEHLIG YNEDIYKKLS DEPSDIIPL FETAITQVAK RISILSRAQS ANNNDKDPEN TSMDTDSLLL NSLPTFQLIL NSNANQIPLR DLDSEHVSKI V RLSGIIIS TSVLSSRATY LSIMCRNCRH TTSITINNFN SITGNTVSLP RSCLSTIESE SSMANESNIG DESTKKNCGP DP YIIIHES SKFIDQQFLK LQEIPELVPV GEMPRNLTMT CDRYLTNKVI PGTRVTIVGI YSIYNSKNGA GSGRSGGGNG GSG VAIRTP YIKILGIQSD VETSSIWNSV TMFTEEEEEE FLQLSRNPKL YEILTNSIAP SIFGNEDIKK AIVCLLMGGS KKIL PDGMR LRGDINVLLL GDPGTAKSQL LKFVEKVSPI AVYTSGKGSS AAGLTASVQR DPMTREFYLE GGAMVLADGG VVCID EFDK MRDEDRVAIH EAMEQQTISI AKAGITTVLN SRTSVLAAAN PIYGRYDDLK SPGDNIDFQT TILSRFDMIF IVKDDH NEE RDISIANHVI NIHTGNANAM QNQQEENGSE ISIEKMKRYI TYCRLKCAPR LSPQAAEKLS SNFVTIRKQL LINELES TE RSSIPITIRQ LEAIIRITES LAKLELSPIA QERHVDEAIR LFQASTMDAA SQDPIGGLNQ ASGTSLSEIR RFEQELKR R LPIGWSTSYQ TLRREFVDTH RFSQLALDKA LYALEKHETI QLRHQGQNIY RSGV

UniProtKB: Minichromosome maintenance protein 5

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Macromolecule #5: DNA replication licensing factor MCM6

MacromoleculeName: DNA replication licensing factor MCM6 / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA helicase
Source (natural)Organism: Saccharomyces cerevisiae S288C (yeast) / Strain: ATCC 204508 / S288c
Molecular weightTheoretical: 113.110211 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)
SequenceString: MSSPFPADTP SSNRPSNSSP PPSSIGAGFG SSSGLDSQIG SRLHFPSSSQ PHVSNSQTGP FVNDSTQFSS QRLQTDGSAT NDMEGNEPA RSFKSRALNH VKKVDDVTGE KVREAFEQFL EDFSVQSTDT GEVEKVYRAQ IEFMKIYDLN TIYIDYQHLS M RENGALAM ...String:
MSSPFPADTP SSNRPSNSSP PPSSIGAGFG SSSGLDSQIG SRLHFPSSSQ PHVSNSQTGP FVNDSTQFSS QRLQTDGSAT NDMEGNEPA RSFKSRALNH VKKVDDVTGE KVREAFEQFL EDFSVQSTDT GEVEKVYRAQ IEFMKIYDLN TIYIDYQHLS M RENGALAM AISEQYYRFL PFLQKGLRRV VRKYAPELLN TSDSLKRSEG DEGQADEDEQ QDDDMNGSSL PRDSGSSAAP GN GTSAMAT RSITTSTSPE QTERVFQISF FNLPTVHRIR DIRSEKIGSL LSISGTVTRT SEVRPELYKA SFTCDMCRAI VDN VEQSFK YTEPTFCPNP SCENRAFWTL NVTRSRFLDW QKVRIQENAN EIPTGSMPRT LDVILRGDSV ERAKPGDRCK FTGV EIVVP DVTQLGLPGV KPSSTLDTRG ISKTTEGLNS GVTGLRSLGV RDLTYKISFL ACHVISIGSN IGASSPDANS NNRET ELQM AANLQANNVY QDNERDQEVF LNSLSSDEIN ELKEMVKDEH IYDKLVRSIA PAVFGHEAVK KGILLQMLGG VHKSTV EGI KLRGDINICV VGDPSTSKSQ FLKYVVGFAP RSVYTSGKAS SAAGLTAAVV RDEEGGDYTI EAGALMLADN GICCIDE FD KMDISDQVAI HEAMEQQTIS IAKAGIHATL NARTSILAAA NPVGGRYNRK LSLRGNLNMT APIMSRFDLF FVILDDCN E KIDTELASHI VDLHMKRDEA IEPPFSAEQL RRYIKYARTF KPILTKEARS YLVEKYKELR KDDAQGFSRS SYRITVRQL ESMIRLSEAI ARANCVDEIT PSFIAEAYDL LRQSIIRVDV DDVEMDEEFD NIESQSHAAS GNNDDNDDGT GSGVITSEPP ADIEEGQSE ATARPGTSEK KKTTVTYDKY VSMMNMIVRK IAEVDREGAE ELTAVDIVDW YLLQKENDLG SLAEYWEERR L AFKVIKRL VKDRILMEIH GTRHNLRDLE NEENENNKTV YVIHPNCEVL DQLEPQDSS

UniProtKB: DNA replication licensing factor MCM6

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Macromolecule #6: DNA replication licensing factor MCM7

MacromoleculeName: DNA replication licensing factor MCM7 / type: protein_or_peptide / ID: 6 / Number of copies: 1 / Enantiomer: LEVO / EC number: DNA helicase
Source (natural)Organism: Saccharomyces cerevisiae S288C (yeast) / Strain: ATCC 204508 / S288c
Molecular weightTheoretical: 95.049875 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)
SequenceString: MSAALPSIQL PVDYNNLFNE ITDFLVTFKQ DTLSSDATRN ENEDENLDAE NIEQHLLEKG PKYMAMLQKV ANRELNSVII DLDDILQYQ NEKFLQGTQA DDLVSAIQQN ANHFTELFCR AIDNNMPLPT KEIDYKDDVL DVILNQRRLR NERMLSDRTN E IRSENLMD ...String:
MSAALPSIQL PVDYNNLFNE ITDFLVTFKQ DTLSSDATRN ENEDENLDAE NIEQHLLEKG PKYMAMLQKV ANRELNSVII DLDDILQYQ NEKFLQGTQA DDLVSAIQQN ANHFTELFCR AIDNNMPLPT KEIDYKDDVL DVILNQRRLR NERMLSDRTN E IRSENLMD TTMDPPSSMN DALREVVEDE TELFPPNLTR RYFLYFKPLS QNCARRYRKK AISSKPLSVR QIKGDFLGQL IT VRGIITR VSDVKPAVEV IAYTCDQCGY EVFQEVNSRT FTPLSECTSE ECSQNQTKGQ LFMSTRASKF SAFQECKIQE LSQ QVPVGH IPRSLNIHVN GTLVRSLSPG DIVDVTGIFL PAPYTGFKAL KAGLLTETYL EAQFVRQHKK KFASFSLTSD VEER VMELI TSGDVYNRLA KSIAPEIYGN LDVKKALLLL LVGGVDKRVG DGMKIRGDIN VCLMGDPGVA KSQLLKAICK ISPRG VYTT GKGSSGVGLT AAVMKDPVTD EMILEGGALV LADNGICCID EFDKMDESDR TAIHEVMEQQ TISISKAGIN TTLNAR TSI LAAANPLYGR YNPRLSPLDN INLPAALLSR FDILFLMLDI PSRDDDEKLA EHVTYVHMHN KQPDLDFTPV EPSKMRE YI AYAKTKRPVM SEAVNDYVVQ AYIRLRQDSK REMDSKFSFG QATPRTLLGI IRLSQALAKL RLADMVDIDD VEEALRLV R VSKESLYQET NKSKEDESPT TKIFTIIKKM LQETGKNTLS YENIVKTVRL RGFTMLQLSN CIQEYSYLNV WHLINEGNT LKFVDDGTMD TDQEDSLVST PKLAPQTTAS ANVSAQDSDI DLQDA

UniProtKB: DNA replication licensing factor MCM7

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Macromolecule #7: DNA replication complex GINS protein PSF1

MacromoleculeName: DNA replication complex GINS protein PSF1 / type: protein_or_peptide / ID: 7 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae S288C (yeast) / Strain: ATCC 204508 / S288c
Molecular weightTheoretical: 24.230576 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)
SequenceString: MYGDLGNKLV LEAKRTKQLY ARSNQDVNLP MYHEDIIRNI LKEVSNLRKN TEYLKEQQQL GMLDDKVAKC QYFVTLLCME RNKRCLLAY QRLRTDILDS MAWNNNGLDL MSSITFSQQD TNNLSHQEQE YLKEYCDLIT DLKSGDLVDI DLSGSLVPPS D VFIDVRVL ...String:
MYGDLGNKLV LEAKRTKQLY ARSNQDVNLP MYHEDIIRNI LKEVSNLRKN TEYLKEQQQL GMLDDKVAKC QYFVTLLCME RNKRCLLAY QRLRTDILDS MAWNNNGLDL MSSITFSQQD TNNLSHQEQE YLKEYCDLIT DLKSGDLVDI DLSGSLVPPS D VFIDVRVL KDAGEIQTEY GVFNLIKDSQ FFVRQSDVER LIQQGYLQKI

UniProtKB: DNA replication complex GINS protein PSF1

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Macromolecule #8: DNA replication complex GINS protein PSF2

MacromoleculeName: DNA replication complex GINS protein PSF2 / type: protein_or_peptide / ID: 8 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae S288C (yeast) / Strain: ATCC 204508 / S288c
Molecular weightTheoretical: 25.096807 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)
SequenceString: MSLPAHLQQT FSPEEIQFIV ENEPIKIFPR ITTRQKIRGD DRGTGNHTRW QLITTDDKAL NNMVAMRSTE VVLWIALLLK QQSKCSIVA PQWLTTKELD RKIQYEKTHP DRFSELPWNW LVLARILFNK AKDDFHDPIH ELRGKIQDLR EIRQIKVLKG L KYLNESHL ...String:
MSLPAHLQQT FSPEEIQFIV ENEPIKIFPR ITTRQKIRGD DRGTGNHTRW QLITTDDKAL NNMVAMRSTE VVLWIALLLK QQSKCSIVA PQWLTTKELD RKIQYEKTHP DRFSELPWNW LVLARILFNK AKDDFHDPIH ELRGKIQDLR EIRQIKVLKG L KYLNESHL QLDNLSLLEI NELRPFITEI MDKLREIHTA SLTAGTENDE EEFNI

UniProtKB: DNA replication complex GINS protein PSF2

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Macromolecule #9: DNA replication complex GINS protein PSF3

MacromoleculeName: DNA replication complex GINS protein PSF3 / type: protein_or_peptide / ID: 9 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae S288C (yeast) / Strain: ATCC 204508 / S288c
Molecular weightTheoretical: 21.977135 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)
SequenceString: MGYYDIDDVL ADGTEFPCKF QYDIPGLGYL ENNPGRPITK NTKLSLPLWL ARILAIVGGD EALVDEEPVP FVELLPPDMF STKVMNAIK TDPVALDLHS INSHFFSLAI KWIMLFSEKE LANVVSELLL QRAQELNHHA SSLSIDLNAD STGKNSANTN I ATSTFLLK ...String:
MGYYDIDDVL ADGTEFPCKF QYDIPGLGYL ENNPGRPITK NTKLSLPLWL ARILAIVGGD EALVDEEPVP FVELLPPDMF STKVMNAIK TDPVALDLHS INSHFFSLAI KWIMLFSEKE LANVVSELLL QRAQELNHHA SSLSIDLNAD STGKNSANTN I ATSTFLLK LEEMEKEIYK KSHESYKDTK RWMFKK

UniProtKB: DNA replication complex GINS protein PSF3

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Macromolecule #10: DNA replication complex GINS protein SLD5

MacromoleculeName: DNA replication complex GINS protein SLD5 / type: protein_or_peptide / ID: 10 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae S288C (yeast) / Strain: ATCC 204508 / S288c
Molecular weightTheoretical: 33.983617 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)
SequenceString: MDINIDDILA ELDKETTAVD STKITQGSSS TTHRDANTIV GSSLDLNDKT QIYVSPQQDF SDLMKSWKNE RCSPELLPYP HQLMKRLLN RISMQSQLIE NISMGFLDMQ NASNANPPMP NESKLPLLCM ETELERLKFV IRSYIRCRLS KIDKFSLYLR Q LNEDENSL ...String:
MDINIDDILA ELDKETTAVD STKITQGSSS TTHRDANTIV GSSLDLNDKT QIYVSPQQDF SDLMKSWKNE RCSPELLPYP HQLMKRLLN RISMQSQLIE NISMGFLDMQ NASNANPPMP NESKLPLLCM ETELERLKFV IRSYIRCRLS KIDKFSLYLR Q LNEDENSL ISLTDLLSKD EIKYHDTHSL IWLKLVNDSI LKYMPEELQA INDTEGSVNM IDEPDWNKFV FIHVNGPPDG KW NEDPLLQ ENEFGKPCYT VTIPDLKEEV ELTIGSIYVM RYEVIRDLLR DDKVALI

UniProtKB: DNA replication complex GINS protein SLD5

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Macromolecule #11: Cell division control protein 45

MacromoleculeName: Cell division control protein 45 / type: protein_or_peptide / ID: 11 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae S288C (yeast) / Strain: ATCC 204508 / S288c
Molecular weightTheoretical: 74.324836 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)
SequenceString: MYYGISQFSE AYNKILRNSS SHSSCQLVIF VSCLNIDALC ATKMLSLLFK KQLVQSQIVP IFGYSELRRH YSQLDDNINS LLLVGFGGV IDLEAFLEID PQEYVIDTDE KSGEQSFRRD IYVLDAHRPW NLDNIFGSQI IQCFDDGTVD DTLGEQKEAY Y KLLELDEE ...String:
MYYGISQFSE AYNKILRNSS SHSSCQLVIF VSCLNIDALC ATKMLSLLFK KQLVQSQIVP IFGYSELRRH YSQLDDNINS LLLVGFGGV IDLEAFLEID PQEYVIDTDE KSGEQSFRRD IYVLDAHRPW NLDNIFGSQI IQCFDDGTVD DTLGEQKEAY Y KLLELDEE SGDDELSGDE NDNNGGDDEA TDADEVTDED EEDEDETISN KRGNSSIGPN DLSKRKQRKK QIHEYEGVLE EY YSQGTTV VNSISAQIYS LLSAIGETNL SNLWLNILGT TSLDIAYAQV YNRLYPLLQD EVKRLTPSSR NSVKTPDTLT LNI QPDYYL FLLRHSSLYD SFYYSNYVNA KLSLWNENGK KRLHKMFARM GIPLSTAQET WLYMDHSIKR ELGIIFDKNL DRYG LQDII RDGFVRTLGY RGSISASEFV EALTALLEVG NSTDKDSVKI NNDNNDDTDG EEEEDNSAQK LTNLRKRWVS NFWLS WDAL DDRKVELLNR GIQLAQDLQR AIFNTGVAIL EKKLIKHLRI YRLCVLQDGP DLDLYRNPLT LLRLGNWLIE CCAESE DKQ LLPMVLASID ENTDTYLVAG LTPRYPRGLD TIHTKKPILN NFSMAFQQIT AETDAKVRID NFESSIIEIR REDLSPF LE KLTLSGLL

UniProtKB: Cell division control protein 45

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Macromolecule #12: DNA polymerase alpha-binding protein

MacromoleculeName: DNA polymerase alpha-binding protein / type: protein_or_peptide / ID: 12 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae S288C (yeast) / Strain: ATCC 204508 / S288c
Molecular weightTheoretical: 104.543391 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)
SequenceString: MVSVIDKLVF DFGGKTLVSL APDNNTLCVA NKNGLTKILK TNNPEEEPET LDSSKLVSSI KCYSNSHFLM TTMQGDALRY NIDSSQEEL LARFALPLRD CCVIHSGKMA VFGGDDLELI LLELDDETHK KHAIKIDEQV SQISYNSQMN ILAVSMINGK V QIFSLTST ...String:
MVSVIDKLVF DFGGKTLVSL APDNNTLCVA NKNGLTKILK TNNPEEEPET LDSSKLVSSI KCYSNSHFLM TTMQGDALRY NIDSSQEEL LARFALPLRD CCVIHSGKMA VFGGDDLELI LLELDDETHK KHAIKIDEQV SQISYNSQMN ILAVSMINGK V QIFSLTST IPNKVHELND YIVANSYDDT HRDKILSNMM DDIDKDNDND LSETADPDEN NVADPEFCAA NRICTRVAWH PK GLHFALP CADDTVKIFS IKGYSLQKTL STNLSSTKAH FIDLQFDPLR GTYIAAVDLN NKLTVWNWET SEIHYTREFK RKI TNIAWK IQADSKTLDL VLGTWSGSIA IVQNLAESVV SNIPDQSVAE SSTKHGLFVD SESDLENLEG NDDINKSDKL FSDI TQEAN AEDVFTQTHD GPSGLSEKRK YNFEDEEDFI DDDDGAGYIS GKKPHNEHSY SRVHKTHSFP ISLANTGKFR YMPFS PAGT PFGFTDRRYL TMNEVGYVST VKNSEQYSIT VSFFDVGRFR EYHFEDLFGY DLCFLNEKGT LFGQSKTGQI QYRPHD SIH SNWTKIIPLQ AGERITSVAA TPVRVIVGTS LGYFRSFNQF GVPFAVEKTS PIVALTAQNY RVFSVHYSQF HGLSYSL SE LGTSSKRYYK RECPLPMSLP NINSDMKKDA NLDYYNFNPM GIKSLFFSSY GDPCIFGSDN TLLLLSKWRS PEESKWLP I LDSNMEIWKM SGGKETTDIH VWPLALAYDT LNCILVKGKH IWPEFPLPLP SEMEIRMPVF VKSKLLEENK AILNKKNEI GADTEAEEGE EDKEIQIPVS MAAEEEYLRS KVLSELLTDT LENDGEMYGN ENEVLAALNG AYDKALLRLF ASACSDQNVE KALSLAHEL KQDRALTAAV KISERAELPS LVKKINNIRE ARYEQQLK

UniProtKB: DNA polymerase alpha-binding protein

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Macromolecule #14: DNA polymerase epsilon subunit B

MacromoleculeName: DNA polymerase epsilon subunit B / type: protein_or_peptide / ID: 14 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae S288C (yeast) / Strain: ATCC 204508 / S288c
Molecular weightTheoretical: 78.425852 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)
SequenceString: MFGSGNVLPV KIQPPLLRPL AYRVLSRKYG LSIKSDGLSA LAEFVGTNIG ANWRQGPATI KFLEQFAAVW KQQERGLFID QSGVKEVIQ EMKEREKVEW SHEHPIQHEE NILGRTDDDE NNSDDEMPIA ADSSLQNVSL SSPMRQPTER DEYKQPFKPE S SKALDWRD ...String:
MFGSGNVLPV KIQPPLLRPL AYRVLSRKYG LSIKSDGLSA LAEFVGTNIG ANWRQGPATI KFLEQFAAVW KQQERGLFID QSGVKEVIQ EMKEREKVEW SHEHPIQHEE NILGRTDDDE NNSDDEMPIA ADSSLQNVSL SSPMRQPTER DEYKQPFKPE S SKALDWRD YFKVINASQQ QRFSYNPHKM QFIFVPNKKQ NGLGGIAGFL PDIEDKVQMF LTRYYLTNDR VMRNENFQNS DM FNPLSSM VSLQNELSNT NRQQQSSSMS ITPIKNLLGR DAQNFLLLGL LNKNFKGNWS LEDPSGSVEI DISQTIPTQG HYY VPGCMV LVEGIYYSVG NKFHVTSMTL PPGERREITL ETIGNLDLLG IHGISNNNFI ARLDKDLKIR LHLLEKELTD HKFV ILGAN LFLDDLKIMT ALSKILQKLN DDPPTLLIWQ GSFTSVPVFA SMSSRNISSS TQFKNNFDAL ATLLSRFDNL TENTT MIFI PGPNDLWGSM VSLGASGTLP QDPIPSAFTK KINKVCKNVV WSSNPTRIAY LSQEIVIFRD DLSGRFKRHR LEFPFN ESE DVYTENDNMM SKDTDIVPID ELVKEPDQLP QKVQETRKLV KTILDQGHLS PFLDSLRPIS WDLDHTLTLC PIPSTMV LC DTTSAQFDLT YNGCKVINPG SFIHNRRARY MEYVPSSKKT IQEEIYI

UniProtKB: DNA polymerase epsilon subunit B

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Macromolecule #13: DNA (71-mer)

MacromoleculeName: DNA (71-mer) / type: dna / ID: 13 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Molecular weightTheoretical: 22.13815 KDa
SequenceString: (DT)(DA)(DG)(DA)(DG)(DT)(DA)(DG)(DG)(DA) (DA)(DG)(DT)(DG)(DA)(DT)(DG)(DG)(DT)(DA) (DA)(DG)(DT)(DG)(DA)(DT)(DT)(DA)(DG) (DA)(DG)(DA)(DA)(DT)(DT)(DG)(DG)(DA)(DG) (DA) (DG)(DT)(DG)(DT)(DG)(DT) ...String:
(DT)(DA)(DG)(DA)(DG)(DT)(DA)(DG)(DG)(DA) (DA)(DG)(DT)(DG)(DA)(DT)(DG)(DG)(DT)(DA) (DA)(DG)(DT)(DG)(DA)(DT)(DT)(DA)(DG) (DA)(DG)(DA)(DA)(DT)(DT)(DG)(DG)(DA)(DG) (DA) (DG)(DT)(DG)(DT)(DG)(DT)(DT)(DT) (DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT) (DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT)(DT)(DT)

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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: 7.6
Component:
ConcentrationFormulaName
100.0 mMKClsodium potassium
25.0 mMHEPES4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid
4.0 mMMgCl2magnesium chloride
0.5 mMATP-gamma-SAdenosine 5'-O-(3-thio)triphosphate
GridModel: C-flat-1.2/1.3 / Material: GOLD / Mesh: 300
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV
Details: blot with filter paper for 3-4 seconds before plunging..

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 81000
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4092 pixel / Number grids imaged: 4 / Number real images: 21776 / Average exposure time: 4.5 sec. / Average electron dose: 53.0 e/Å2
Details: Images were collected in movie-mode containing 40 frames.
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Software: (Name: cryoSPARC (ver. v3.0.1), cryoSPARC (ver. v2.15.0))
Final angle assignmentType: MAXIMUM LIKELIHOOD
Software: (Name: cryoSPARC (ver. v3.0.1), cryoSPARC (ver. v2.15.0))
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: EXACT BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 7.39 Å / Resolution method: FSC 0.143 CUT-OFF
Software: (Name: cryoSPARC (ver. v3.0.1), cryoSPARC (ver. v2.15.0))
Number images used: 7939

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Atomic model buiding 1

Initial modelPDB ID:

6skl


Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementSpace: REAL / Protocol: AB INITIO MODEL
Output model

PDB-8w7s:
Yeast replisome in state IV

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