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- EMDB-10227: Cryo-EM structure of the CMG Fork Protection Complex at a replica... -

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Basic information

Entry
Database: EMDB / ID: EMD-10227
TitleCryo-EM structure of the CMG Fork Protection Complex at a replication fork - Conformation 1
Map dataConformation1 CMG-Csm3-Tof1-Mrc1-Ctf4 and a DNA fork
Sample
  • Complex: Conformation 1 of CMG-Csm3-Tof1-Mrc1-Ctf4 with a fork DNA
    • Complex: CMG-Csm3-Tof1-Mrc1-Ctf4
      • Protein or peptide: x 14 types
    • Complex: DNA
      • DNA: x 2 types
  • Ligand: x 3 types
Function / homology
Function and homology information


establishment of sister chromatid cohesion / maintenance of DNA repeat elements / Unwinding of DNA / replication fork arrest / Cul8-RING ubiquitin ligase complex / meiotic chromosome segregation / 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 / maintenance of DNA repeat elements / Unwinding of DNA / replication fork arrest / Cul8-RING ubiquitin ligase complex / meiotic chromosome segregation / 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 / establishment of mitotic sister chromatid cohesion / DNA replication checkpoint signaling / 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 / single-stranded DNA helicase activity / replication fork protection complex / mitotic DNA replication initiation / silent mating-type cassette heterochromatin formation / regulation of DNA-templated DNA replication initiation / DNA strand elongation involved in DNA replication / mitotic sister chromatid cohesion / replication fork processing / nuclear chromosome / DNA unwinding involved in DNA replication / nuclear replication fork / DNA replication origin binding / subtelomeric heterochromatin formation / DNA replication initiation / heterochromatin formation / DNA helicase activity / meiotic cell cycle / helicase activity / DNA-templated DNA replication / nucleosome assembly / single-stranded DNA binding / mitotic cell cycle / DNA helicase / DNA replication / chromosome, telomeric region / DNA repair / DNA damage response / chromatin binding / ATP hydrolysis activity / DNA binding / nucleoplasm / ATP binding / identical protein binding / metal ion binding / nucleus / cytosol / cytoplasm
Similarity search - Function
Chromosome segregation in meiosis protein 3 / TIPIN/Csm3/Swi3 / Replication Fork Protection Component Swi3 / Timeless / Timeless, N-terminal / Timeless protein / : / DNA polymerase alpha-binding protein Ctf4, C-terminal domain / Minichromosome loss protein Mcl1, middle region / Minichromosome loss protein, Mcl1, middle region ...Chromosome segregation in meiosis protein 3 / TIPIN/Csm3/Swi3 / Replication Fork Protection Component Swi3 / Timeless / Timeless, N-terminal / Timeless protein / : / 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 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 MCM3 / DNA replication licensing factor MCM2 / 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 / Topoisomerase 1-associated factor 1 / DNA polymerase alpha-binding protein / DNA replication complex GINS protein SLD5 ...DNA replication licensing factor MCM3 / DNA replication licensing factor MCM2 / 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 / Topoisomerase 1-associated factor 1 / DNA polymerase alpha-binding protein / DNA replication complex GINS protein SLD5 / Chromosome segregation in meiosis protein 3 / Cell division control protein 45 / DNA replication complex GINS protein PSF3 / DNA replication complex GINS protein PSF1
Similarity search - Component
Biological speciesSaccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) / synthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.7 Å
AuthorsYeeles J / Baretic D / Jenkyn-Bedford M
Funding support United Kingdom, 1 items
OrganizationGrant numberCountry
Medical Research Council (MRC, United Kingdom)MC_UP_1201/12 United Kingdom
CitationJournal: Mol Cell / Year: 2020
Title: Cryo-EM Structure of the Fork Protection Complex Bound to CMG at a Replication Fork.
Authors: Domagoj Baretić / Michael Jenkyn-Bedford / Valentina Aria / Giuseppe Cannone / Mark Skehel / Joseph T P Yeeles /
Abstract: The eukaryotic replisome, organized around the Cdc45-MCM-GINS (CMG) helicase, orchestrates chromosome replication. Multiple factors associate directly with CMG, including Ctf4 and the heterotrimeric ...The eukaryotic replisome, organized around the Cdc45-MCM-GINS (CMG) helicase, orchestrates chromosome replication. Multiple factors associate directly with CMG, including Ctf4 and the heterotrimeric fork protection complex (Csm3/Tof1 and Mrc1), which has important roles including aiding normal replication rates and stabilizing stalled forks. How these proteins interface with CMG to execute these functions is poorly understood. Here we present 3 to 3.5 Å resolution electron cryomicroscopy (cryo-EM) structures comprising CMG, Ctf4, and the fork protection complex at a replication fork. The structures provide high-resolution views of CMG-DNA interactions, revealing a mechanism for strand separation, and show Csm3/Tof1 "grip" duplex DNA ahead of CMG via a network of interactions important for efficient replication fork pausing. Although Mrc1 was not resolved in our structures, we determine its topology in the replisome by cross-linking mass spectrometry. Collectively, our work reveals how four highly conserved replisome components collaborate with CMG to facilitate replisome progression and maintain genome stability.
History
DepositionAug 16, 2019-
Header (metadata) releaseMay 6, 2020-
Map releaseMay 6, 2020-
UpdateJun 17, 2020-
Current statusJun 17, 2020Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.018
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.018
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6skl
  • Surface level: 0.018
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_10227.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationConformation1 CMG-Csm3-Tof1-Mrc1-Ctf4 and a DNA fork
Voxel sizeX=Y=Z: 1.049 Å
Density
Contour LevelBy AUTHOR: 0.018 / Movie #1: 0.018
Minimum - Maximum-0.033392858 - 0.06839484
Average (Standard dev.)0.0000379676 (±0.00300201)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions360360360
Spacing360360360
CellA=B=C: 377.64 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.0491.0491.049
M x/y/z360360360
origin x/y/z0.0000.0000.000
length x/y/z377.640377.640377.640
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS360360360
D min/max/mean-0.0330.0680.000

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Supplemental data

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Sample components

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Entire : Conformation 1 of CMG-Csm3-Tof1-Mrc1-Ctf4 with a fork DNA

EntireName: Conformation 1 of CMG-Csm3-Tof1-Mrc1-Ctf4 with a fork DNA
Components
  • Complex: Conformation 1 of CMG-Csm3-Tof1-Mrc1-Ctf4 with a fork DNA
    • Complex: CMG-Csm3-Tof1-Mrc1-Ctf4
      • 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
      • Protein or peptide: Topoisomerase 1-associated factor 1
      • Protein or peptide: Chromosome segregation in meiosis protein 3
    • Complex: DNA
      • DNA: DNA fork, leading-strand template
      • DNA: DNA fork, lagging-strand template
  • Ligand: ZINC ION
  • Ligand: PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER
  • Ligand: MAGNESIUM ION

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Supramolecule #1: Conformation 1 of CMG-Csm3-Tof1-Mrc1-Ctf4 with a fork DNA

SupramoleculeName: Conformation 1 of CMG-Csm3-Tof1-Mrc1-Ctf4 with a fork DNA
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#16
Molecular weightTheoretical: 1.4 MDa

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Supramolecule #2: CMG-Csm3-Tof1-Mrc1-Ctf4

SupramoleculeName: CMG-Csm3-Tof1-Mrc1-Ctf4 / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1-#12, #15-#16
Source (natural)Organism: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)

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Supramolecule #3: DNA

SupramoleculeName: DNA / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #13-#14
Source (natural)Organism: synthetic construct (others)
Recombinant expressionOrganism: synthetic construct (others)

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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 / EC number: DNA helicase
Source (natural)Organism: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (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

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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 (strain ATCC 204508 / S288c) (yeast)
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

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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 (strain ATCC 204508 / S288c) (yeast)
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

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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 (strain ATCC 204508 / S288c) (yeast)
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

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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 (strain ATCC 204508 / S288c) (yeast)
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

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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 (strain ATCC 204508 / S288c) (yeast)
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

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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 (strain ATCC 204508 / S288c) (yeast)
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

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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 (strain ATCC 204508 / S288c) (yeast)
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

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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 / Details: His-tag at the N-terminus (24 residues) / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Molecular weightTheoretical: 24.437859 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)
SequenceString: MGSSHHHHHH SSGLVPRGSH MASMGYYDID DVLADGTEFP CKFQYDIPGL GYLENNPGRP ITKNTKLSLP LWLARILAIV GGDEALVDE EPVPFVELLP PDMFSTKVMN AIKTDPVALD LHSINSHFFS LAIKWIMLFS EKELANVVSE LLLQRAQELN H HASSLSID ...String:
MGSSHHHHHH SSGLVPRGSH MASMGYYDID DVLADGTEFP CKFQYDIPGL GYLENNPGRP ITKNTKLSLP LWLARILAIV GGDEALVDE EPVPFVELLP PDMFSTKVMN AIKTDPVALD LHSINSHFFS LAIKWIMLFS EKELANVVSE LLLQRAQELN H HASSLSID LNADSTGKNS ANTNIATSTF LLKLEEMEKE IYKKSHESYK DTKRWMFKK

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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 (strain ATCC 204508 / S288c) (yeast)
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

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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 (strain ATCC 204508 / S288c) (yeast)
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

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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 (strain ATCC 204508 / S288c) (yeast)
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

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Macromolecule #15: Topoisomerase 1-associated factor 1

MacromoleculeName: Topoisomerase 1-associated factor 1 / type: protein_or_peptide / ID: 15 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Molecular weightTheoretical: 141.296875 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)
SequenceString: MSADLQQGTT NAADFSLTVL RARIALLATA IGGPDYTSQI DPPPYKLGDD CLACLKDLKR WFKLVDDQQK RWDVAMAVAE YRILTDDLL PILIDWENKC SLAAKLAKNN PDHEEFRNKA YYDKIALNCL QLLVLMTWPL IVTEQSSSNQ ITLYGELKKH Q LVYKKTIL ...String:
MSADLQQGTT NAADFSLTVL RARIALLATA IGGPDYTSQI DPPPYKLGDD CLACLKDLKR WFKLVDDQQK RWDVAMAVAE YRILTDDLL PILIDWENKC SLAAKLAKNN PDHEEFRNKA YYDKIALNCL QLLVLMTWPL IVTEQSSSNQ ITLYGELKKH Q LVYKKTIL SMESGKVLRA AIRLALDVIK IDRLSRTPRD NMVLKLVLNF FRNVIAIEPG EFTINTKKSM PKKGITSIDT LP PNVSMDD ISLNTVISSF HKNKVFGFLL TLTSSLSKEF DQDFINIPLL EIMFYFTKDV NQELLFPRQF ETGTHSKVVN KNE SSSANN IVTSAGFELS KLLQKEHQMR KNVIKHTSAR HSRFGGLLSI QTPDKTRLTV SGSQALVDEK IALQKLDDSK KWNK RIIKK HQSVAAEGLP NSLLNSQTGK AIFFTESNGK HFKEFINNFI DSGFNILLHS VTNYFTTEQD RMVTLEQVEY LLFFA WFVK YQLLRSKIDN SADIKQVSEA LKEVTFILVS SLLRSAYDLK NWTVTHAGMI AFNELLNLVS RTKAAQEEDS TDIEFI VSR LFSDERIQLL SNLPKIGSKY SLQFMKSCIE LTHSVLKVLE QYSDDKTLVI EGKSRRQKKF NISEGDITKL IEEENVD RD EALDILTSSL RSIEVNFQKV QANYMTEPVI ETYINFLERF RELEDDSIKK VFSFFHRVFV QAKEQALLFR FDLIILLR E MLSPDGLDRM SRSRKYVSQF SDYFLARLKK RLKKSPAWFV GLLFPPLHNS EVGFYQRYGE YNVLNNESMY AAPASQFKP IPDEEALPPS ILLDMKYGVL VSTLLDDGKT ELLDQLLKHI THTLDIFKSW LTVNVNAGKE TVNPPNEYFT LTGVLNNDPI FKDKDYRAL LLLIGYSIPR KINEPCFLPG TVEVSDLTVS CELVKKYLST PFETPNGLPS SSYLLRVRSE KDSFSHNEQD G WEGDDDYD YNDPYIVPDD QILSKSDAAY FKDLDNNASD KLKGTKFSKG IARSKKKDKR KRRKGEAKTN LPMFGDQDDE RP QTVRERH GVFSKEFISD SEDDEDLMNP IFFENETYMR WLLDKNNGQL TEDRYIQFAK FAAERMNNGG VVTGDYTSLF GGS IPSIES IRATESSSFA PDKSLISLAS HVASEMSIFD VNNNNNNQLS DDDVNSESRN SLGSSQPSNS QNMFQSEVYS RKES TKRSL EASAADESDE DEEAIRLFGK KSRVVLSQGD SDD

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Macromolecule #16: Chromosome segregation in meiosis protein 3

MacromoleculeName: Chromosome segregation in meiosis protein 3 / type: protein_or_peptide / ID: 16 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Molecular weightTheoretical: 36.40259 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)
SequenceString: MDQDFDSLLL GFNDSDSVQK DPTVPNGLDG SVVDPTIADP TAITARKRRP QVKLTAEKLL SDKGLPYVLK NAHKRIRISS KKNSYDNLS NIIQFYQLWA HELFPKAKFK DFMKICQTVG KTDPVLREYR VSLFRDEMGM SFDVGTRETG QDLERQSPMV E EHVTSAEE ...String:
MDQDFDSLLL GFNDSDSVQK DPTVPNGLDG SVVDPTIADP TAITARKRRP QVKLTAEKLL SDKGLPYVLK NAHKRIRISS KKNSYDNLS NIIQFYQLWA HELFPKAKFK DFMKICQTVG KTDPVLREYR VSLFRDEMGM SFDVGTRETG QDLERQSPMV E EHVTSAEE RPIVADSFAQ DKRNVNNVDY DNDEDDDIYH LSYRNRRGRV LDERGNNETV LNNVVPPKED LDALLKTFRV QG PVGLEEN EKKLLLGWLD AHRKMEKGSM TEEDVQLIQS LEEWEMNDIE GQHTHYDLLP GGDEFGVDQD ELDAMKEMGF

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Macromolecule #13: DNA fork, leading-strand template

MacromoleculeName: DNA fork, leading-strand template / type: dna / ID: 13
Details: Cy3-label at 5'-end. Five phosphorothioate backbone linkages between residues at the very 3'-end. The dT 16 residues from the 5'-end was biotinylated.
Number of copies: 1 / Classification: DNA
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 26.396836 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)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT)(DT) (DT)(DT)(DT)(DT)(DT)

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Macromolecule #14: DNA fork, lagging-strand template

MacromoleculeName: DNA fork, lagging-strand template / type: dna / ID: 14 / Details: dT (residue 47) biotinylated. / Number of copies: 1 / Classification: DNA
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 18.524887 KDa
SequenceString: (DG)(DG)(DC)(DA)(DG)(DG)(DC)(DA)(DG)(DG) (DC)(DA)(DG)(DG)(DC)(DA)(DC)(DA)(DC)(DA) (DC)(DT)(DC)(DT)(DC)(DC)(DA)(DA)(DT) (DT)(DC)(DT)(DC)(DT)(DA)(DA)(DT)(DC)(DA) (DC) (DT)(DT)(DA)(DC)(DC)(DA) ...String:
(DG)(DG)(DC)(DA)(DG)(DG)(DC)(DA)(DG)(DG) (DC)(DA)(DG)(DG)(DC)(DA)(DC)(DA)(DC)(DA) (DC)(DT)(DC)(DT)(DC)(DC)(DA)(DA)(DT) (DT)(DC)(DT)(DC)(DT)(DA)(DA)(DT)(DC)(DA) (DC) (DT)(DT)(DA)(DC)(DC)(DA)(DT)(DC) (DA)(DC)(DT)(DT)(DC)(DC)(DT)(DA)(DC)(DT) (DC)(DT) (DA)

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Macromolecule #17: ZINC ION

MacromoleculeName: ZINC ION / type: ligand / ID: 17 / Number of copies: 5 / Formula: ZN
Molecular weightTheoretical: 65.409 Da

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Macromolecule #18: PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER

MacromoleculeName: PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER / type: ligand / ID: 18 / Number of copies: 3 / Formula: ANP
Molecular weightTheoretical: 506.196 Da
Chemical component information

ChemComp-ANP:
PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER / AMP-PNP, energy-carrying molecule analogue*YM

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Macromolecule #19: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 19 / Number of copies: 3 / Formula: MG
Molecular weightTheoretical: 24.305 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

Concentration0.5 mg/mL
BufferpH: 7.5
Component:
ConcentrationFormulaName
25.0 mMHEPES4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid
150.0 mMNaOAcsodium acetate
0.5 mMTCEPtris(2-carboxyethyl)phosphine
0.1 mMAMP-PNPadenylyl-imidodiphosphate
0.7 mMMg(OAc)2magnesium acetate
0.005 %TWEEN20Polyoxyethylene (20) sorbitan monolaurate
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Support film - Film thickness: 5.0 nm / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR / Details: 15 mA
VitrificationCryogen name: ETHANE / Chamber humidity: 80 % / Chamber temperature: 277.15 K / Instrument: HOMEMADE PLUNGER
Details: Three microlitres of sample was applied on a grid and incubated for 15-30 s at 4 degC before manually blotting with filter paper for 10 s and plunge-freezing in liquid ethane..
DetailsIn vitro reconstitution from individual components: CMG, Csm3-Tof1, Mrc1, Ctf4 and a DNA fork

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Calibrated defocus max: 3.0 µm / Calibrated defocus min: 0.5 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.6 µm / Nominal defocus min: 1.4 µm / Nominal magnification: 130000
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 K2 QUANTUM (4k x 4k) / Detector mode: COUNTING / Digitization - Frames/image: 1-20 / Number grids imaged: 2 / Number real images: 6682 / Average exposure time: 7.0 sec. / Average electron dose: 37.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 632000
CTF correctionSoftware - Name: Gctf (ver. 0.50)
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0.6)
Final 3D classificationSoftware - Name: RELION (ver. 3.0.6)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0.6)
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.0.6) / Number images used: 35000
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial model(PDB ID:
,
,
)
RefinementSpace: REAL / Protocol: OTHER / Overall B value: 20 / Target criteria: FSC 0.5
Output model

PDB-6skl:
Cryo-EM structure of the CMG Fork Protection Complex at a replication fork - Conformation 1

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