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- EMDB-36064: Cryo-EM structure of Asfv topoisomerase 2 - apo conformer IIa -

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Basic information

Entry
Database: EMDB / ID: EMD-36064
TitleCryo-EM structure of Asfv topoisomerase 2 - apo conformer IIa
Map data
Sample
  • Organelle or cellular component: ASFV Topoisomerase 2
    • Protein or peptide: DNA topoisomerase 2
KeywordsTopoisomerase / ASFV / VIRAL PROTEIN / ISOMERASE
Function / homology
Function and homology information


sister chromatid segregation / DNA topoisomerase type II (double strand cut, ATP-hydrolyzing) activity / DNA topoisomerase (ATP-hydrolysing) / DNA topological change / DNA binding / ATP binding / metal ion binding / cytoplasm
Similarity search - Function
C-terminal associated domain of TOPRIM / C-terminal associated domain of TOPRIM / DNA topoisomerase II, eukaryotic-type / : / DNA topoisomerase, type IIA, alpha-helical domain superfamily / DNA topoisomerase, type IIA, domain A / DNA topoisomerase, type IIA, domain A, alpha-beta / DNA gyrase/topoisomerase IV, subunit A / DNA Topoisomerase IV / DNA topoisomerase, type IIA ...C-terminal associated domain of TOPRIM / C-terminal associated domain of TOPRIM / DNA topoisomerase II, eukaryotic-type / : / DNA topoisomerase, type IIA, alpha-helical domain superfamily / DNA topoisomerase, type IIA, domain A / DNA topoisomerase, type IIA, domain A, alpha-beta / DNA gyrase/topoisomerase IV, subunit A / DNA Topoisomerase IV / DNA topoisomerase, type IIA / DNA topoisomerase, type IIA, conserved site / DNA topoisomerase II signature. / TopoisomeraseII / DNA topoisomerase, type IIA, subunit B, C-terminal / DNA topoisomerase, type IIA-like domain superfamily / Histidine kinase/HSP90-like ATPase superfamily / Ribosomal protein S5 domain 2-type fold, subgroup
Similarity search - Domain/homology
Biological speciesAfrican swine fever virus
Methodsingle particle reconstruction / cryo EM / Resolution: 2.31 Å
AuthorsChang C-W / Tsai M-D
Funding support Taiwan, 1 items
OrganizationGrant numberCountry
Academia Sinica (Taiwan)AS-KPQ-109-TPP2 Taiwan
CitationJournal: Commun Chem / Year: 2024
Title: A unified view on enzyme catalysis by cryo-EM study of a DNA topoisomerase.
Authors: Chiung-Wen Mary Chang / Shun-Chang Wang / Chun-Hsiung Wang / Allan H Pang / Cheng-Han Yang / Yao-Kai Chang / Wen-Jin Wu / Ming-Daw Tsai /
Abstract: The theories for substrate recognition in enzyme catalysis have evolved from lock-key to induced fit, then conformational selection, and conformational selection followed by induced fit. However, the ...The theories for substrate recognition in enzyme catalysis have evolved from lock-key to induced fit, then conformational selection, and conformational selection followed by induced fit. However, the prevalence and consensus of these theories require further examination. Here we use cryogenic electron microscopy and African swine fever virus type 2 topoisomerase (AsfvTop2) to demonstrate substrate binding theories in a joint and ordered manner: catalytic selection by the enzyme, conformational selection by the substrates, then induced fit. The apo-AsfvTop2 pre-exists in six conformers that comply with the two-gate mechanism directing DNA passage and release in the Top2 catalytic cycle. The structures of AsfvTop2-DNA-inhibitor complexes show that substantial induced-fit changes occur locally from the closed apo-conformer that however is too far-fetched for the open apo-conformer. Furthermore, the ATPase domain of AsfvTop2 in the MgAMP-PNP-bound crystal structures coexist in reduced and oxidized forms involving a disulfide bond, which can regulate the AsfvTop2 function.
History
DepositionMay 1, 2023-
Header (metadata) releaseFeb 7, 2024-
Map releaseFeb 7, 2024-
UpdateApr 17, 2024-
Current statusApr 17, 2024Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_36064.png

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Map

FileDownload / File: emd_36064.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

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AxesZ (Sec.)Y (Row.)X (Col.)
0.83 Å/pix.
x 400 pix.
= 332. Å		0.83 Å/pix.
x 400 pix.
= 332. Å		0.83 Å/pix.
x 400 pix.
= 332. Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.83 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.073
Minimum - Maximum-0.18915771 - 0.5287828
Average (Standard dev.)0.000094461524 (±0.010809708)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions400400400
Spacing400400400
CellA=B=C: 332.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_36064_half_map_1.map
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Half map: #1

Fileemd_36064_half_map_2.map
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Sample components

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Entire : ASFV Topoisomerase 2

EntireName: ASFV Topoisomerase 2
Components
  • Organelle or cellular component: ASFV Topoisomerase 2
    • Protein or peptide: DNA topoisomerase 2

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Supramolecule #1: ASFV Topoisomerase 2

SupramoleculeName: ASFV Topoisomerase 2 / type: organelle_or_cellular_component / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: African swine fever virus

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Macromolecule #1: DNA topoisomerase 2

MacromoleculeName: DNA topoisomerase 2 / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: African swine fever virus
Molecular weightTheoretical: 136.422578 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)
SequenceString: MEAFEISDFK EHAKKKSMWA GALNKVTISG LMGVFTEDED LMALPIHRDH CPALLKIFDE LIVNATDHER ACHSKTKKVT YIKISFDKG VFSCENDGPG IPIAKHEQAS LIAKRDVYVP EVASCFFLAG TNINKAKDCI KGGTNGVGLK LAMVHSQWAI L TTADGAQK ...String:
MEAFEISDFK EHAKKKSMWA GALNKVTISG LMGVFTEDED LMALPIHRDH CPALLKIFDE LIVNATDHER ACHSKTKKVT YIKISFDKG VFSCENDGPG IPIAKHEQAS LIAKRDVYVP EVASCFFLAG TNINKAKDCI KGGTNGVGLK LAMVHSQWAI L TTADGAQK YVQQINQRLD IIEPPTITPS REMFTRIELM PVYQELGYAE PLSETEQADL SAWIYLRACQ CAAYVGKGTT IY YNDKPCR TGSVMALAKM YTLLSAPNST IHTATIKADA KPYSLHPLQV AAVVSPKFKK FEHVSIINGV NCVKGEHVTF LKK TINEMV IKKFQQTIKD KNRKTTLRDS CSNIFVVIVG SIPGIEWTGQ RKDELSIAEN VFKTHYSIPS SFLTSMTRSI VDIL LQSIS KKDNHKQVDV DKYTRARNAG GKRAQDCMLL AAEGDSALSL LRTGLTLGKS NPSGPSFDFC GMISLGGVIM NACKK VTNI TTDSGETIMV RNEQLTNNKV LQGIVQVLGL DFNCHYKTQE ERAKLRYGCI VACVDQDLDG CGKILGLLLA YFHLFW PQL IIHGFVKRLL TPLIRVYEKG KTMPVEFYYE QEFDAWAKKQ TSLVNHTVKY YKGLAAHDTH EVKSMFKHFD NMVYTFT LD DSAKELFHIY FGGESELRKR ELCTGVVPLT ETQTQSIHSV RRIPCSLHLQ VDTKAYKLDA IERQIPNFLD GMTRARRK I LAGGVKCFAS NNRERKVFQF GGYVADHMFY HHGDMSLNTS IIKAAQYYPG SSHLYPVFIG IGSFGSRHLG GKDAGSPRY ISVQLASEFI KTMFPAEDSW LLPYVFEDGQ RAEPEYYVPV LPLAIMEYGA NPSEGWKYTT WARQLEDILA LVRAYVDKDN PKHELLHYA IKHKITILPL RPSNYNFKGH LKRFGQYYYS YGTYDISEQR NIITITELPL RVPTVAYIES IKKSSNRMTF I EEIIDYSS SETIEILVKL KPNSLNRIVE EFKETEEQDS IENFLRLRNC LHSHLNFVKP KGGIIEFNSY YEILYAWLPY RR ELYQKRL MREHAVLKLR IIMETAIVRY INESAELNLS HYEDEKEASR ILSEHGFPPL NHTLIISPEF ASIEELNQKA LQG CYTYIL SLQARELLIA AKTRRVEKIK KMQARLDKVE QLLQESPFPG ASVWLEEIDA VEKAIIKGRN TQWKFHHHHH H

UniProtKB: DNA topoisomerase 2

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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: 7.5
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 1 / Number real images: 8900 / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.5 µm
Sample stageCooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER / Details: Robetta prediction server
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.31 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 68014
Initial angle assignmentType: ANGULAR RECONSTITUTION
Final angle assignmentType: ANGULAR RECONSTITUTION
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelChain - Source name: RoseTTAFold / Chain - Initial model type: in silico model
RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-8j89:
Cryo-EM structure of Asfv topoisomerase 2 - apo conformer IIa

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