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- EMDB-27842: Structure of Lates calcarifer Twinkle helicase with ATP and DNA -

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Basic information

Entry
Database: EMDB / ID: EMD-27842
TitleStructure of Lates calcarifer Twinkle helicase with ATP and DNA
Map data
Sample
  • Complex: Twinkle protein, mitochondrial + DNA
    • Protein or peptide: Twinkle mtDNA helicase
    • DNA: DNA (5'-D(P*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*T)-3')
  • Ligand: ADENOSINE-5'-TRIPHOSPHATE
  • Ligand: MAGNESIUM ION
KeywordsHelicase Mitochondrion / REPLICATION-DNA complex
Function / homology
Function and homology information


mitochondrial DNA replication / DNA 5'-3' helicase / mitochondrial nucleoid / single-stranded DNA binding / 5'-3' DNA helicase activity / mitochondrial inner membrane / hydrolase activity / lipid binding / ATP binding
Similarity search - Function
AAA domain / Twinkle-like protein / Archaeal primase DnaG/twinkle-like, TOPRIM domain / DNA helicase, DnaB-like, C-terminal / Superfamily 4 helicase domain profile. / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Twinkle mtDNA helicase
Similarity search - Component
Biological speciesLates calcarifer (barramundi perch) / synthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.51 Å
AuthorsGao Y / Li Z
Funding support United States, 2 items
OrganizationGrant numberCountry
Cancer Prevention and Research Institute of Texas (CPRIT)RR190046 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM142722 United States
CitationJournal: Nucleic Acids Res / Year: 2022
Title: Structural and dynamic basis of DNA capture and translocation by mitochondrial Twinkle helicase.
Authors: Zhuo Li / Parminder Kaur / Chen-Yu Lo / Neil Chopra / Jamie Smith / Hong Wang / Yang Gao /
Abstract: Twinkle is a mitochondrial replicative helicase which can self-load onto and unwind mitochondrial DNA. Nearly 60 mutations on Twinkle have been linked to human mitochondrial diseases. Using cryo- ...Twinkle is a mitochondrial replicative helicase which can self-load onto and unwind mitochondrial DNA. Nearly 60 mutations on Twinkle have been linked to human mitochondrial diseases. Using cryo-electron microscopy (cryo-EM) and high-speed atomic force microscopy (HS-AFM), we obtained the atomic-resolution structure of a vertebrate Twinkle homolog with DNA and captured in real-time how Twinkle is self-loaded onto DNA. Our data highlight the important role of the non-catalytic N-terminal domain of Twinkle. The N-terminal domain directly contacts the C-terminal helicase domain, and the contact interface is a hotspot for disease-related mutations. Mutations at the interface destabilize Twinkle hexamer and reduce helicase activity. With HS-AFM, we observed that a highly dynamic Twinkle domain, which is likely to be the N-terminal domain, can protrude ∼5 nm to transiently capture nearby DNA and initialize Twinkle loading onto DNA. Moreover, structural analysis and subunit doping experiments suggest that Twinkle hydrolyzes ATP stochastically, which is distinct from related helicases from bacteriophages.
History
DepositionAug 15, 2022-
Header (metadata) releaseNov 2, 2022-
Map releaseNov 2, 2022-
UpdateMay 28, 2025-
Current statusMay 28, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_27842.png

Downloads & links

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Map

FileDownload / File: emd_27842.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.)
1.07 Å/pix.
x 320 pix.
= 342.4 Å		1.07 Å/pix.
x 320 pix.
= 342.4 Å		1.07 Å/pix.
x 320 pix.
= 342.4 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1.07 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.15
Minimum - Maximum-0.9868265 - 1.814308
Average (Standard dev.)0.0016533687 (±0.04755441)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 342.40002 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_27842_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_27842_half_map_2.map
Projections & Slices
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Histogram

Histogram (log scale)

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Sample components

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Entire : Twinkle protein, mitochondrial + DNA

EntireName: Twinkle protein, mitochondrial + DNA
Components
  • Complex: Twinkle protein, mitochondrial + DNA
    • Protein or peptide: Twinkle mtDNA helicase
    • DNA: DNA (5'-D(P*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*T)-3')
  • Ligand: ADENOSINE-5'-TRIPHOSPHATE
  • Ligand: MAGNESIUM ION

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Supramolecule #1: Twinkle protein, mitochondrial + DNA

SupramoleculeName: Twinkle protein, mitochondrial + DNA / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Source (natural)Organism: Lates calcarifer (barramundi perch)

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Macromolecule #1: Twinkle mtDNA helicase

MacromoleculeName: Twinkle mtDNA helicase / type: protein_or_peptide / ID: 1 / Number of copies: 6 / Enantiomer: LEVO
Source (natural)Organism: Lates calcarifer (barramundi perch)
Molecular weightTheoretical: 61.29816 KDa
Recombinant expressionOrganism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
SequenceString: MQKEEQDFLS PHVMLGYPES LDEQEEGERE LREVQRIWSS AVPFNDLPED EAQLIKTMFQ ITKVSNATLK KFGVRLFKPT KSLVFPWFA GPDSSLKGLK LLSAQNTDTE KVTYNEATVP KISSYYNLFG LTLVGRMDSE VVLTGHELDT LAVSQATGLP S VALPRGVS ...String:
MQKEEQDFLS PHVMLGYPES LDEQEEGERE LREVQRIWSS AVPFNDLPED EAQLIKTMFQ ITKVSNATLK KFGVRLFKPT KSLVFPWFA GPDSSLKGLK LLSAQNTDTE KVTYNEATVP KISSYYNLFG LTLVGRMDSE VVLTGHELDT LAVSQATGLP S VALPRGVS CLPPMLLPYL EQFKRVTLWL GHDIRSWEAS KIFSRKLGLR RCSLVRPGED RPCPLEALAR GKNLSRIIKT SI PAAHKSI VSFKQLREDV YGELLNTEQV AGVKWTRFPE LNRILKGHRK GELTVFTGPT GSGKTTFISE VALDLCIQGV NTL WGSFQI NNVRLAKIML TQFAMQRLEE NLEQYDFWAD KFEELPLYFM TFHGQQNIKT VLDTMQHAVY LYDINHVIID NLQF MMGQE NLSIDKYAVQ DHIIGAFRKF ATNTSCHVTL IIHPRKEEDD RELQTASIFG SAKASQEADN VLILQEKKLV TCPGR RSLQ VTKNRFDGDV GIFPLDFIKS SLTFSAPIKG KVKLRKVSTK PENEEVGGER GGSEEGRG

UniProtKB: Twinkle mtDNA helicase

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Macromolecule #2: DNA (5'-D(P*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*T)-3')

MacromoleculeName: DNA (5'-D(P*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*TP*T)-3') / type: dna / ID: 2 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 4.517935 KDa
SequenceString:
(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT)(DT)(DT)(DT)

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Macromolecule #3: ADENOSINE-5'-TRIPHOSPHATE

MacromoleculeName: ADENOSINE-5'-TRIPHOSPHATE / type: ligand / ID: 3 / Number of copies: 5 / Formula: ATP
Molecular weightTheoretical: 507.181 Da
Chemical component information

ChemComp-ATP:
ADENOSINE-5'-TRIPHOSPHATE / ATP, energy-carrying molecule*YM

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Macromolecule #4: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 4 / Number of copies: 5 / 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: 8
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 298 K / Instrument: FEI VITROBOT MARK I
Details50 mM Tris (pH 8.0), 150 mM KCl, 3 mM DTT, 1 mM ATP, and 2 mM MgCl2

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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: 0.6 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionSoftware - Name: Gctf / Type: NONE
Startup modelType of model: NONE
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.51 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 44814
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: AB INITIO MODEL
Output model

PDB-8e2l:
Structure of Lates calcarifer Twinkle helicase with ATP and DNA

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