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- EMDB-40237: Structure of wild-type MCM8/9 heterohexamer complex with ATP-gamma-S -

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Basic information

Entry
Database: EMDB / ID: EMD-40237
TitleStructure of wild-type MCM8/9 heterohexamer complex with ATP-gamma-S
Map data
Sample
  • Complex: Structure of wild-type MCM8/9 heterohexamer complex with ATP-gamma-S
    • Protein or peptide: MCM8_HUMAN
    • Protein or peptide: MCM9_HUMAN
KeywordsDNA Replication / DNA repair / DNA BINDING PROTEIN
Function / homology
Function and homology information


MutLbeta complex binding / MutSbeta complex binding / recombinational interstrand cross-link repair / MCM8-MCM9 complex / male gamete generation / mismatch repair involved in maintenance of fidelity involved in DNA-dependent DNA replication / CDC6 association with the ORC:origin complex / female gamete generation / E2F-enabled inhibition of pre-replication complex formation / MutSalpha complex binding ...MutLbeta complex binding / MutSbeta complex binding / recombinational interstrand cross-link repair / MCM8-MCM9 complex / male gamete generation / mismatch repair involved in maintenance of fidelity involved in DNA-dependent DNA replication / CDC6 association with the ORC:origin complex / female gamete generation / E2F-enabled inhibition of pre-replication complex formation / MutSalpha complex binding / Unwinding of DNA / MCM complex / Activation of the pre-replicative complex / Activation of ATR in response to replication stress / DNA helicase activity / protein localization to chromatin / double-strand break repair via homologous recombination / Orc1 removal from chromatin / single-stranded DNA binding / chromosome / DNA helicase / forked DNA-dependent helicase activity / single-stranded 3'-5' DNA helicase activity / four-way junction helicase activity / double-stranded DNA helicase activity / protein stabilization / DNA damage response / chromatin binding / protein-containing complex binding / enzyme binding / ATP hydrolysis activity / nucleoplasm / ATP binding / nucleus
Similarity search - Function
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 / Nucleic acid-binding, OB-fold ...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 / Nucleic acid-binding, OB-fold / ATPases associated with a variety of cellular activities / AAA+ ATPase domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
DNA helicase MCM9 / DNA helicase MCM8
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 6.5 Å
AuthorsLi C / Gao Y
Funding support United States, 1 items
OrganizationGrant numberCountry
Cancer Prevention and Research Institute of Texas (CPRIT)RR190046 United States
CitationJournal: Nucleic Acids Res / Year: 2023
Title: Activity, substrate preference and structure of the HsMCM8/9 helicase.
Authors: David R McKinzey / Chuxuan Li / Yang Gao / Michael A Trakselis /
Abstract: The minichromosomal maintenance proteins, MCM8 and MCM9, are more recent evolutionary additions to the MCM family, only cooccurring in selected higher eukaryotes. Mutations in these genes are ...The minichromosomal maintenance proteins, MCM8 and MCM9, are more recent evolutionary additions to the MCM family, only cooccurring in selected higher eukaryotes. Mutations in these genes are directly linked to ovarian insufficiency, infertility, and several cancers. MCM8/9 appears to have ancillary roles in fork progression and recombination of broken replication forks. However, the biochemical activity, specificities and structures have not been adequately illustrated, making mechanistic determination difficult. Here, we show that human MCM8/9 (HsMCM8/9) is an ATP dependent DNA helicase that unwinds fork DNA substrates with a 3'-5' polarity. High affinity ssDNA binding occurs in the presence of nucleoside triphosphates, while ATP hydrolysis weakens the interaction with DNA. The cryo-EM structure of the HsMCM8/9 heterohexamer was solved at 4.3 Å revealing a trimer of heterodimer configuration with two types of interfacial AAA+ nucleotide binding sites that become more organized upon binding ADP. Local refinements of the N or C-terminal domains (NTD or CTD) improved the resolution to 3.9 or 4.1 Å, respectively, and shows a large displacement in the CTD. Changes in AAA+ CTD upon nucleotide binding and a large swing between the NTD and CTD likely implies that MCM8/9 utilizes a sequential subunit translocation mechanism for DNA unwinding.
History
DepositionMar 27, 2023-
Header (metadata) releaseJun 7, 2023-
Map releaseJun 7, 2023-
UpdateAug 30, 2023-
Current statusAug 30, 2023Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_40237.png

Downloads & links

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Map

FileDownload / File: emd_40237.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.)
1.11 Å/pix.
x 320 pix.
= 355.2 Å		1.11 Å/pix.
x 320 pix.
= 355.2 Å		1.11 Å/pix.
x 320 pix.
= 355.2 Å

Surface

Projections

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1.11 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.05
Minimum - Maximum-0.1469628 - 0.29243407
Average (Standard dev.)-0.0011576707 (±0.012363651)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 355.2 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_40237_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_40237_half_map_2.map
Projections & Slices
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Histogram (log scale)

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Sample components

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Entire : Structure of wild-type MCM8/9 heterohexamer complex with ATP-gamma-S

EntireName: Structure of wild-type MCM8/9 heterohexamer complex with ATP-gamma-S
Components
  • Complex: Structure of wild-type MCM8/9 heterohexamer complex with ATP-gamma-S
    • Protein or peptide: MCM8_HUMAN
    • Protein or peptide: MCM9_HUMAN

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Supramolecule #1: Structure of wild-type MCM8/9 heterohexamer complex with ATP-gamma-S

SupramoleculeName: Structure of wild-type MCM8/9 heterohexamer complex with ATP-gamma-S
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 500 KDa

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Macromolecule #1: MCM8_HUMAN

MacromoleculeName: MCM8_HUMAN / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)
SequenceString: MNGEYRGRGF GRGRFQSWKR GRGGGNFSGK WREREHRPDL SKTTGKRTSE QTPQFLLSTK TPQSMQSTLD RFIPYKGWKL YFSEVYSDSS PLIEKIQAFE KFFTRHIDLY DKDEIERKGS ILVDFKELTE GGEVTNLIPD IATELRDAPE KTLACMGLAI HQVLTKDLER ...String:
MNGEYRGRGF GRGRFQSWKR GRGGGNFSGK WREREHRPDL SKTTGKRTSE QTPQFLLSTK TPQSMQSTLD RFIPYKGWKL YFSEVYSDSS PLIEKIQAFE KFFTRHIDLY DKDEIERKGS ILVDFKELTE GGEVTNLIPD IATELRDAPE KTLACMGLAI HQVLTKDLER HAAELQAQEG LSNDGETMVN VPHIHARVYN YEPLTQLKNV RANYYGKYIA LRGTVVRVSN IKPLCTKMAF LCAACGEIQS FPLPDGKYSL PTKCPVPVCR GRSFTALRSS PLTVTMDWQS IKIQELMSDD QREAGRIPRT IECELVHDLV DSCVPGDTVT ITGIVKVSNA EEGSRNKNDK CMFLLYIEAN SISNSKGQKT KSSEDGCKHG MLMEFSLKDL YAIQEIQAEE NLFKLIVNSL CPVIFGHELV KAGLALALFG GSQKYADDKN RIPIRGDPHI LVVGDPGLGK SQMLQAACNV APRGVYVCGN TTTTSGLTVT LSKDSSSGDF ALEAGALVLG DQGICGIDEF DKMGNQHQAL LEAMEQQSIS LAKAGVVCSL PARTSIIAAA NPVGGHYNKA KTVSENLKMG SALLSRFDLV FILLDTPNEH HDHLLSEHVI AIRAGKQRTI SSATVARMNS QDSNTSVLEV VSEKPLSERL KVVPGETIDP IPHQLLRKYI GYARQYVYPR LSTEAARVLQ DFYLELRKQS QRLNSSPITT RQLESLIRLT EARARLELRE EATKEDAEDI VEIMKYSMLG TYSDEFGNLD FERSQHGSGM SNRSTAKRFI SALNNVAERT YNNIFQFHQL RQIAKELNIQ VADFENFIGS LNDQGYLLKK GPKVYQLQTM

UniProtKB: DNA helicase MCM8

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Macromolecule #2: MCM9_HUMAN

MacromoleculeName: MCM9_HUMAN / type: protein_or_peptide / ID: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)
SequenceString: MNSDQVTLVG QVFESYVSEY HKNDILLILK ERDEDAHYPV VVNAMTLFET NMEIGEYFNM FPSEVLTIFD SALRRSALTI LQSLSQPEAV SMKQNLHARI SGLPVCPELV REHIPKTKDV GHFLSVTGTV IRTSLVKVLE FERDYMCNKC KHVFVIKADF EQYYTFCRPS ...String:
MNSDQVTLVG QVFESYVSEY HKNDILLILK ERDEDAHYPV VVNAMTLFET NMEIGEYFNM FPSEVLTIFD SALRRSALTI LQSLSQPEAV SMKQNLHARI SGLPVCPELV REHIPKTKDV GHFLSVTGTV IRTSLVKVLE FERDYMCNKC KHVFVIKADF EQYYTFCRPS SCPSLESCDS SKFTCLSGLS SSPTRCRDYQ EIKIQEQVQR LSVGSIPRSM KVILEDDLVD SCKSGDDLTI YGIVMQRWKP FQQDVRCEVE IVLKANYIQV NNEQSSGIIM DEEVQKEFED FWEYYKSDPF AGRNVILASL CPQVFGMYLV KLAVAMVLAG GIQRTDATGT RVRGESHLLL VGDPGTGKSQ FLKYAAKITP RSVLTTGIGS TSAGLTVTAV KDSGEWNLEA GALVLADAGL CCIDEFNSLK EHDRTSIHEA MEQQTISVAK AGLVCKLNTR TTILAATNPK GQYDPQESVS VNIALGSPLL SRFDLILVLL DTKNEDWDRI ISSFILENKG YPSKSEKLWS MEKMKTYFCL IRNLQPTLSD VGNQVLLRYY QMQRQSDCRN AARTTIRLLE SLIRLAEAHA RLMFRDTVTL EDAITVVSVM ESSMQGGALL GGVNALHTSF PENPGEQYQR QCELILEKLE LQSLLSEELR RLERLQNQSV HQSQPRVLEV ETTPGSLRNG PGEESNFRTS SQQEINYSTH IFSPGGSPEG SPVLDPPPHL EPNRSTSRKH SAQHKNNRDD SLDWFDFMAT HQSEPKNTVV VSPHPKTSGE NMASKISNST SQGKEKSEPG QRSKVDIGLL PSPGETGVPW RADNVESNKK KRLALDSEAA VSADKPDSVL THHVPRNLQK LCKERAQKLC RNSTRVPAQC TVPSHPQSTP VHSPDRMLDS PKRKRPKSLA QVEEPAIENV KPPGSPVAKL AKFTFKQKSK LIHSFEDHSH VSPGATKIAV HSPKISQRRT RRDAALPVKR PGKLTSTPGN QISSQPQGET KEVSQQPPEK HGPREKVMCA PEKRIIQPEL ELGNETGCAH LTCEGDKKEE VSGSNKSGKV HACTLARLAN FCFTPPSESK SKSPPPERKN RGERGPSSPP TTTAPMRVSK RKSFQLRGST EKLIVSKESL FTLPELGDEA FDCDWDEEMR KKS

UniProtKB: DNA helicase MCM9

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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: 8
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 298 K / Instrument: FEI VITROBOT MARK I

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 49.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 3.0 µm / Nominal defocus min: 0.7000000000000001 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 6.5 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 57384
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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