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- EMDB-27977: SsoMCM hexamer bound to Mg/ADP-BeFx and 16-mer oligo-dT. Class 2 -

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Basic information

Entry
Database: EMDB / ID: EMD-27977
TitleSsoMCM hexamer bound to Mg/ADP-BeFx and 16-mer oligo-dT. Class 2
Map dataunsharpened map
Sample
  • Complex: MCM hexamer bound to Mg/ADP-BeFx and 16-mer oligo-dT. Class 2
    • Protein or peptide: Minichromosome maintenance protein MCM
    • DNA: 16-mer oligo-dT
  • Ligand: ZINC ION
  • Ligand: MAGNESIUM ION
  • Ligand: [[[(2R,3S,4R,5R)-5-(6-aminopurin-9-yl)-3,4-bis(oxidanyl)oxolan-2-yl]methoxy-oxidanyl-phosphoryl]oxy-oxidanyl-phosphoryl]oxy-tris(fluoranyl)beryllium
  • Ligand: water
KeywordsHelicase / ATPase / REPLICATION / TRANSFERASE-DNA complex
Function / homology
Function and homology information


MCM complex / DNA duplex unwinding / helicase activity / single-stranded DNA binding / DNA helicase / DNA replication / ATP hydrolysis activity / ATP binding / identical protein binding
Similarity search - Function
: / MCM protein C-terminal winged helix-turn-helix domain / 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 protein C-terminal winged helix-turn-helix domain / 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 / 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
Minichromosome maintenance protein MCM
Similarity search - Component
Biological speciesSaccharolobus solfataricus P2 (archaea) / synthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.76 Å
AuthorsMeagher M / Myasnikov A / Enemark EJ
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM136313 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM098771 United States
CitationJournal: Int J Mol Sci / Year: 2022
Title: Two Distinct Modes of DNA Binding by an MCM Helicase Enable DNA Translocation.
Authors: Martin Meagher / Alexander Myasnikov / Eric J Enemark /
Abstract: A six-subunit ATPase ring forms the central hub of the replication forks in all domains of life. This ring performs a helicase function to separate the two complementary DNA strands to be replicated ...A six-subunit ATPase ring forms the central hub of the replication forks in all domains of life. This ring performs a helicase function to separate the two complementary DNA strands to be replicated and drives the replication machinery along the DNA. Disruption of this helicase/ATPase ring is associated with genetic instability and diseases such as cancer. The helicase/ATPase rings of eukaryotes and archaea consist of six minichromosome maintenance (MCM) proteins. Prior structural studies have shown that MCM rings bind one encircled strand of DNA in a spiral staircase, suggesting that the ring pulls this strand of DNA through its central pore in a hand-over-hand mechanism where the subunit at the bottom of the staircase dissociates from DNA and re-binds DNA one step above the staircase. With high-resolution cryo-EM, we show that the MCM ring of the archaeal organism binds an encircled DNA strand in two different modes with different numbers of subunits engaged to DNA, illustrating a plausible mechanism for the alternating steps of DNA dissociation and re-association that occur during DNA translocation.
History
DepositionAug 29, 2022-
Header (metadata) releaseDec 14, 2022-
Map releaseDec 14, 2022-
UpdateJun 19, 2024-
Current statusJun 19, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_27977.png

Downloads & links

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Map

FileDownload / File: emd_27977.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationunsharpened map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.08 Å/pix.
x 256 pix.
= 276.48 Å		1.08 Å/pix.
x 256 pix.
= 276.48 Å		1.08 Å/pix.
x 256 pix.
= 276.48 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.08 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.5
Minimum - Maximum-0.31100762 - 2.124089
Average (Standard dev.)0.007946384 (±0.09122916)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 276.48 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_27977_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: sharpened map

Fileemd_27977_additional_1.map
Annotationsharpened map
Projections & Slices
AxesZYX

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Density Histograms

Histogram

Histogram (log scale)

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Half map: half-map A

Fileemd_27977_half_map_1.map
Annotationhalf-map A
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half-map B

Fileemd_27977_half_map_2.map
Annotationhalf-map B
Projections & Slices
AxesZYX

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Sample components

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Entire : MCM hexamer bound to Mg/ADP-BeFx and 16-mer oligo-dT. Class 2

EntireName: MCM hexamer bound to Mg/ADP-BeFx and 16-mer oligo-dT. Class 2
Components
  • Complex: MCM hexamer bound to Mg/ADP-BeFx and 16-mer oligo-dT. Class 2
    • Protein or peptide: Minichromosome maintenance protein MCM
    • DNA: 16-mer oligo-dT
  • Ligand: ZINC ION
  • Ligand: MAGNESIUM ION
  • Ligand: [[[(2R,3S,4R,5R)-5-(6-aminopurin-9-yl)-3,4-bis(oxidanyl)oxolan-2-yl]methoxy-oxidanyl-phosphoryl]oxy-oxidanyl-phosphoryl]oxy-tris(fluoranyl)beryllium
  • Ligand: water

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Supramolecule #1: MCM hexamer bound to Mg/ADP-BeFx and 16-mer oligo-dT. Class 2

SupramoleculeName: MCM hexamer bound to Mg/ADP-BeFx and 16-mer oligo-dT. Class 2
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Source (natural)Organism: Saccharolobus solfataricus P2 (archaea)

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Macromolecule #1: Minichromosome maintenance protein MCM

MacromoleculeName: Minichromosome maintenance protein MCM / type: protein_or_peptide / ID: 1 / Number of copies: 6 / Enantiomer: LEVO / EC number: DNA helicase
Source (natural)Organism: Saccharolobus solfataricus P2 (archaea) / Strain: ATCC 35092 / DSM 1617 / JCM 11322 / P2
Molecular weightTheoretical: 68.641961 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: SLEIPSKQID YRDVFIEFLT TFKGNNNQNK YIERINELVA YRKKSLIIEF SDVLSFNENL AYEIINNTKI ILPILEGALY DHILQLDPT YQRDIEKVHV RIVGIPRVIE LRKIRSTDIG KLITIDGILV KVTPVKERIY KATYKHIHPD CMQEFEWPED E EMPEVLEM ...String:
SLEIPSKQID YRDVFIEFLT TFKGNNNQNK YIERINELVA YRKKSLIIEF SDVLSFNENL AYEIINNTKI ILPILEGALY DHILQLDPT YQRDIEKVHV RIVGIPRVIE LRKIRSTDIG KLITIDGILV KVTPVKERIY KATYKHIHPD CMQEFEWPED E EMPEVLEM PTICPKCGKP GQFRLIPEKT KLIDWQKAVI QERPEEVPSG QLPRQLEIIL EDDLVDSARP GDRVKVTGIL DI KQDSPVK RGSRAVFDIY MKVSSIEVSG GSGGSSEEDE KKIKDLAKDP WIRDRIISSI APSIYGHWEL KEALALALFG GVP KVLEDT RIRGDIHILI IGDPGTAKSQ MLQFISRVAP RAVYTTGKGS TAAGLTAAVV REKGTGEYYL EAGALVLADG GIAV IDEID KMRDEDRVAI HEAMEQQTVS IAKAGIVAKL NARAAVIAAG NPKFGRYISE RPVSDNINLP PTILSRFDLI FILKD QPGE QDRELANYIL DVHSGKSTKN IIDIDTLRKY IAYARKYVTP KITSEAKNLI TDFFVEMRKK SSETPDSPIL ITPRQL EAL IRISEAYAKM ALKAEVTRED AERAINIMRL FLESVGVDME SGKIDID

UniProtKB: Minichromosome maintenance protein MCM

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Macromolecule #2: 16-mer oligo-dT

MacromoleculeName: 16-mer oligo-dT / type: dna / ID: 2 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 4.822127 KDa
SequenceString:
(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT)(DT)(DT)(DT)(DT)

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Macromolecule #3: ZINC ION

MacromoleculeName: ZINC ION / type: ligand / ID: 3 / Number of copies: 6 / Formula: ZN
Molecular weightTheoretical: 65.409 Da

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Macromolecule #4: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 4 / Number of copies: 4 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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Macromolecule #5: [[[(2R,3S,4R,5R)-5-(6-aminopurin-9-yl)-3,4-bis(oxidanyl)oxolan-2-...

MacromoleculeName: [[[(2R,3S,4R,5R)-5-(6-aminopurin-9-yl)-3,4-bis(oxidanyl)oxolan-2-yl]methoxy-oxidanyl-phosphoryl]oxy-oxidanyl-phosphoryl]oxy-tris(fluoranyl)beryllium
type: ligand / ID: 5 / Number of copies: 4 / Formula: 08T
Molecular weightTheoretical: 492.201 Da
Chemical component information

ChemComp-08T:
[[[(2R,3S,4R,5R)-5-(6-aminopurin-9-yl)-3,4-bis(oxidanyl)oxolan-2-yl]methoxy-oxidanyl-phosphoryl]oxy-oxidanyl-phosphoryl]oxy-tris(fluoranyl)beryllium

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Macromolecule #6: water

MacromoleculeName: water / type: ligand / ID: 6 / Number of copies: 12 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
WATER

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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
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 78.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 1.8 µm / Nominal defocus min: 0.8 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionDetails: Particles were selected from a subset of micrographs with cryoSPARC blob picker. 2D class averages were calculated, selected and used as templates with cryoSPARC template picker.
Startup modelType of model: NONE / Details: An ab initio model was generated by cryoSPARC
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 2.76 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.3.1) / Software - details: Homo_Refine_New / Number images used: 776823
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 2.14.2)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.3.1) / Software - details: Homo_Refine_New
Final 3D classificationNumber classes: 2 / Software - Name: cryoSPARC (ver. 3.3.1) / Software - details: Hetero_Refine
Details: Two major classes of structure with 16-mer oligo-dT are observed. This entry reports the second class.
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

6mii


Chain - Chain ID: B / Chain - Source name: PDB / Chain - Initial model type: experimental model
DetailsBond distance and angle restraints for a tetrahedral Zn2+ coordination were applied. Bond distance and angle restraints for a octahedral Mg2+ coordination were applied.
RefinementSpace: REAL / Protocol: OTHER
Output model

PDB-8eai:
SsoMCM hexamer bound to Mg/ADP-BeFx and 16-mer oligo-dT. Class 2

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