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- EMDB-43755: BsaXI-DNA complex II -

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Basic information

Entry
Database: EMDB / ID: EMD-43755
TitleBsaXI-DNA complex II
Map dataBsaXI-DNA complex conformation P40-J521 map file P40-J524
Sample
  • Complex: Quaternary complex of RM, S and DNA complex
    • Protein or peptide: RM.BsaXI
    • Protein or peptide: S.BsaXI
    • DNA: DNA (41-MER)
    • DNA: DNA (41-MER)
  • Ligand: S-ADENOSYL-L-HOMOCYSTEINE
  • Ligand: S-ADENOSYLMETHIONINE
Keywordsrestriction nuclease / restriction-modification systems / Type IIB / R-M complex / DNA BINDING PROTEIN / DNA BINDING PROTEIN-DNA complex
Function / homology: / Type I restriction modification DNA specificity domain / Type I restriction modification DNA specificity domain superfamily / Type I restriction modification DNA specificity domain / DNA restriction-modification system / DNA binding / Uncharacterized protein / :
Function and homology information
Biological speciesGeobacillus stearothermophilus (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.06 Å
AuthorsShen BW / Stoddard BL / Xu S
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)Ro1-GM105691 United States
CitationJournal: To Be Published
Title: Structural and biochemical analysis of subunit assembly, DNA recognition and cleavage by a Type IIB restriction-modification enzyme: BsaXI
Authors: Shen BW / Heiter D / Xu S / Stoddard BL
History
DepositionFeb 21, 2024-
Header (metadata) releaseOct 30, 2024-
Map releaseOct 30, 2024-
UpdateOct 30, 2024-
Current statusOct 30, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_43755.png

Downloads & links

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Map

FileDownload / File: emd_43755.map.gz / Format: CCP4 / Size: 137.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationBsaXI-DNA complex conformation P40-J521 map file P40-J524
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.12 Å/pix.
x 330 pix.
= 369.6 Å		1.12 Å/pix.
x 330 pix.
= 369.6 Å		1.12 Å/pix.
x 330 pix.
= 369.6 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.12 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.08
Minimum - Maximum-0.35610756 - 0.8353333
Average (Standard dev.)0.00048428625 (±0.02029813)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions330330330
Spacing330330330
CellA=B=C: 369.6 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_43755_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: BsaXI-DNA complex conformation J521 P40-J524 half map B

Fileemd_43755_half_map_1.map
AnnotationBsaXI-DNA complex conformation J521 P40-J524 half map B
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: BsaXI DNA complex conformation J521 p40-J524 half map A

Fileemd_43755_half_map_2.map
AnnotationBsaXI_DNA complex conformation J521 p40-J524 half map A
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Quaternary complex of RM, S and DNA complex

EntireName: Quaternary complex of RM, S and DNA complex
Components
  • Complex: Quaternary complex of RM, S and DNA complex
    • Protein or peptide: RM.BsaXI
    • Protein or peptide: S.BsaXI
    • DNA: DNA (41-MER)
    • DNA: DNA (41-MER)
  • Ligand: S-ADENOSYL-L-HOMOCYSTEINE
  • Ligand: S-ADENOSYLMETHIONINE

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Supramolecule #1: Quaternary complex of RM, S and DNA complex

SupramoleculeName: Quaternary complex of RM, S and DNA complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#4
Source (natural)Organism: Geobacillus stearothermophilus (bacteria) / Strain: Cpw230
Molecular weightTheoretical: 2.69 kDa/nm

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Macromolecule #1: RM.BsaXI

MacromoleculeName: RM.BsaXI / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
EC number: site-specific DNA-methyltransferase (adenine-specific)
Source (natural)Organism: Geobacillus stearothermophilus (bacteria) / Strain: Cpw230
Molecular weightTheoretical: 107.195234 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MKNWQRIVEA KLEQQKHKVA EISLENGTVN YSKKIKHNRN LKALTGDEEI VRAFLIDRLV NELDYKPEYL ETEKEYTIKG GHSKINPRV DVLVKDDKGN PFFFIEVKAP NKFEEDKDEI EGQLFALAQA EERDFKTKVK YLVYYTVELI DDEIVDRAII I DFEKYPTY ...String:
MKNWQRIVEA KLEQQKHKVA EISLENGTVN YSKKIKHNRN LKALTGDEEI VRAFLIDRLV NELDYKPEYL ETEKEYTIKG GHSKINPRV DVLVKDDKGN PFFFIEVKAP NKFEEDKDEI EGQLFALAQA EERDFKTKVK YLVYYTVELI DDEIVDRAII I DFEKYPTY TDWSNGGFIS TGTELTAGYG EPKKQPLIKG HEKYDLRVRI DREEIEGLGR NLHNVLWGGG GTNDSEIFYS LV NIILAKI QDEYEKEDGQ EYDFQVYQYG DNVESPQKLF DRINALYKRA LREQLNVTDE QKIAEDNVIN RNKFPLNKLV YTV QALESL SFLEGRNSLD GKDILGDFFE SIIRDGFKQT KGQFFTPTPI VKFILYALQL DKLAIDRLNN DRELPLIIDP SAGS GTFLI EAMKLITKEV KYKQNHKVKS SRQITKRFEE LFMPDHNENK WAREYLYGCE INFDLGTASK VNMILHGDGS ANIFV QDGL LPFRFYVKET SPNYLETASP DALYGDKEVN GKFDVVVSNP PFSVDLDTQT QREVRNAFLF GDKKNSENLF IERYYQ LLK EGGRLGVVLP ESVFDTTENK YIRLFIFKYF KVKAVVSLPQ VTFEPFTSTK TSLLFAQKKT KEEVEQWNEL WDKYGKE WS LLKTRINDYF SYFVKGRPLN KKWAPDVVKD IQEGNEDNIR KNIFRFLKDH IKEEDKNLEI KDLLIKYAEE ISSISKHE K ETDVFGFYNA WWVFGEVAKE LDYPIFMAEA ENVGYKRTKK GEKPMPNDLY DLEYAPSTLD CEKVLSSFDI EINALEASK TKLSVEKGLL EEKLKDKEDK ENEKIQKRLN KISELLETIE NQLDSIRSKK LEVEGILEKY YENNKLKEEY SERDDEELIN HFKHGVLYQ YRSEDILLRN KTVHKILDEI RQGVIWD

UniProtKB: UNIPROTKB: A0A4D7QEP1

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Macromolecule #2: S.BsaXI

MacromoleculeName: S.BsaXI / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Geobacillus stearothermophilus (bacteria) / Strain: Cpw230
Molecular weightTheoretical: 55.038824 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MGLIQRRNFS TFASEPSVRF DFNYMKSVTP TTEEYYTYKS LFEVVPSTVP TLDESEPFKY AEIGHVSKNG EVFPVTLSFE DRDELNEDL FKKIEKGDIF LPERGNILIS AIRPYLNKIV LIKEDDKTDI YFTKAFIQIK PLINSRILYY ALRTIFSEKI N AVSRQGKG ...String:
MGLIQRRNFS TFASEPSVRF DFNYMKSVTP TTEEYYTYKS LFEVVPSTVP TLDESEPFKY AEIGHVSKNG EVFPVTLSFE DRDELNEDL FKKIEKGDIF LPERGNILIS AIRPYLNKIV LIKEDDKTDI YFTKAFIQIK PLINSRILYY ALRTIFSEKI N AVSRQGKG YPTLKEDDLK TIQFSKKVID NLLAKEEELI SNIDALEKDI KELKSIQRSK KEIVDEVFSS HFNINMVELM AL DSQRRVD VGLSSISSLN STIRYSYRWN KMKLIQKYLY RDIDCIEPLG KYILSSNNGW SPESVVGGEG IPILGQEHLE FDG VLNVSP TKATTKTKNN MENFFIQEGD LFISRGNTVD LVGLACVVET EVTEDIIYPD LYIRLKIDEK VIHKKYLALL FNSF FGRLY FKYVSKGKNQ TMVKISSNEL LNYYLPIPPM EEQLEIVGKI EEQIGAQNEI EKQIEEKRNQ IRVIIEETAR S

UniProtKB: Uncharacterized protein

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Macromolecule #3: DNA (41-MER)

MacromoleculeName: DNA (41-MER) / type: dna / ID: 3 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Geobacillus stearothermophilus (bacteria)
Molecular weightTheoretical: 16.097412 KDa
SequenceString: (DA)(DA)(DT)(DA)(DA)(DG)(DC)(DT)(DG)(DA) (DA)(DT)(DA)(DT)(DT)(DG)(DT)(DC)(DG)(DG) (DA)(6MA)(DC)(DC)(DA)(DA)(DG)(DT)(DC) (DT)(DC)(DC)(DA)(DT)(DA)(DT)(DG)(DG)(DA) (DA)(DT)(DT)(DA)(DA)(DT)(DA) ...String:
(DA)(DA)(DT)(DA)(DA)(DG)(DC)(DT)(DG)(DA) (DA)(DT)(DA)(DT)(DT)(DG)(DT)(DC)(DG)(DG) (DA)(6MA)(DC)(DC)(DA)(DA)(DG)(DT)(DC) (DT)(DC)(DC)(DA)(DT)(DA)(DT)(DG)(DG)(DA) (DA)(DT)(DT)(DA)(DA)(DT)(DA)(DA)(DG) (DC)(DT)(DA)(DG)

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Macromolecule #4: DNA (41-MER)

MacromoleculeName: DNA (41-MER) / type: dna / ID: 4 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Geobacillus stearothermophilus (bacteria)
Molecular weightTheoretical: 15.949256 KDa
SequenceString: (DC)(DT)(DA)(DG)(DC)(DT)(DT)(DA)(DT)(DT) (DA)(DA)(DT)(DT)(DC)(DC)(DA)(DT)(DA)(DT) (DG)(DG)(DA)(DG)(DA)(DC)(DT)(DT)(DG) (DG)(DT)(DT)(DC)(DC)(DG)(DA)(DC)(DA)(DA) (DT) (DA)(DT)(DT)(DC)(DA)(DG) ...String:
(DC)(DT)(DA)(DG)(DC)(DT)(DT)(DA)(DT)(DT) (DA)(DA)(DT)(DT)(DC)(DC)(DA)(DT)(DA)(DT) (DG)(DG)(DA)(DG)(DA)(DC)(DT)(DT)(DG) (DG)(DT)(DT)(DC)(DC)(DG)(DA)(DC)(DA)(DA) (DT) (DA)(DT)(DT)(DC)(DA)(DG)(DC)(DT) (DT)(DA)(DT)(DT)

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Macromolecule #5: S-ADENOSYL-L-HOMOCYSTEINE

MacromoleculeName: S-ADENOSYL-L-HOMOCYSTEINE / type: ligand / ID: 5 / Number of copies: 1 / Formula: SAH
Molecular weightTheoretical: 384.411 Da
Chemical component information

ChemComp-SAH:
S-ADENOSYL-L-HOMOCYSTEINE

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Macromolecule #6: S-ADENOSYLMETHIONINE

MacromoleculeName: S-ADENOSYLMETHIONINE / type: ligand / ID: 6 / Number of copies: 1 / Formula: SAM
Molecular weightTheoretical: 398.437 Da
Chemical component information

ChemComp-SAM:
S-ADENOSYLMETHIONINE

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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

Concentration0.3 mg/mL
BufferpH: 8 / Component - Concentration: 20.0 mM / Component - Name: TrisHCl / Details: 20 mM TrisHCl, 250 mM Nacl, 2 mM CaCl2
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 278 K / Instrument: FEI VITROBOT MARK IV
DetailsSample was monodisperse.

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Electron microscopy #1

Microscopy ID1
MicroscopeFEI TALOS ARCTICA
TemperatureMin: 170.0 K
Specialist opticsPhase plate: VOLTA PHASE PLATE
Image recordingImage recording ID: 1 / Film or detector model: DIRECT ELECTRON DE-10 (5k x 4k) / Detector mode: COUNTING / Digitization - Frames/image: 2-50 / Number grids imaged: 3 / Number real images: 1579 / Average exposure time: 2.0 sec. / Average electron dose: 40.0 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Calibrated defocus max: 5.0 µm / Calibrated defocus min: 1.2 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 5.0 µm / Nominal defocus min: 1.2 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Electron microscopy #1~

Microscopy ID1
MicroscopeFEI TITAN KRIOS
TemperatureMin: 170.0 K / Max: 170.0 K
Specialist opticsPhase plate: OTHER
Image recordingImage recording ID: 2 / Film or detector model: DIRECT ELECTRON DE-20 (5k x 3k) / Detector mode: COUNTING / Digitization - Frames/image: 2-50 / Number grids imaged: 1 / Number real images: 2238 / Average exposure time: 2.0 sec. / Average electron dose: 40.0 e/Å2
Electron beamAcceleration voltage: 3000 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 5.0 µm / Nominal defocus min: 1.2 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Image recording ID1
Startup modelType of model: NONE
Final reconstructionNumber classes used: 4 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.06 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.3) / Number images used: 244448
Initial angle assignmentType: COMMON LINE / Software - Name: cryoSPARC (ver. 3.3)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.3)
Final 3D classificationNumber classes: 2 / Software - Name: cryoSPARC (ver. 3.3)
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelChain - Source name: Other / Chain - Initial model type: experimental model / Details: manual built model in coot
RefinementSpace: REAL / Protocol: FLEXIBLE FIT
Output model

PDB-8w2q:
BsaXI-DNA complex II

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