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- EMDB-15520: Cryo-EM structure of human tankyrase 2 SAM-PARP filament (G1032W ... -

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Basic information

Entry
Database: EMDB / ID: EMD-15520
TitleCryo-EM structure of human tankyrase 2 SAM-PARP filament (G1032W mutant)
Map data
Sample
  • Complex: TNKS2 SAM-PARP (867-1162) filament
    • Protein or peptide: Poly [ADP-ribose] polymerase tankyrase-2
  • Ligand: ZINC ION
Function / homology
Function and homology information


XAV939 stabilizes AXIN / NAD+ ADP-ribosyltransferase / protein localization to chromosome, telomeric region / negative regulation of telomere maintenance via telomere lengthening / protein auto-ADP-ribosylation / protein poly-ADP-ribosylation / pericentriolar material / NAD+-protein ADP-ribosyltransferase activity / positive regulation of telomere capping / NAD+ ADP-ribosyltransferase activity ...XAV939 stabilizes AXIN / NAD+ ADP-ribosyltransferase / protein localization to chromosome, telomeric region / negative regulation of telomere maintenance via telomere lengthening / protein auto-ADP-ribosylation / protein poly-ADP-ribosylation / pericentriolar material / NAD+-protein ADP-ribosyltransferase activity / positive regulation of telomere capping / NAD+ ADP-ribosyltransferase activity / Transferases; Glycosyltransferases; Pentosyltransferases / positive regulation of telomere maintenance via telomerase / nucleotidyltransferase activity / TCF dependent signaling in response to WNT / Degradation of AXIN / Wnt signaling pathway / Regulation of PTEN stability and activity / protein polyubiquitination / positive regulation of canonical Wnt signaling pathway / nuclear envelope / chromosome, telomeric region / Ub-specific processing proteases / Golgi membrane / perinuclear region of cytoplasm / enzyme binding / metal ion binding / nucleus / cytosol / cytoplasm
Similarity search - Function
Ankyrin repeat / Ankyrin repeats (many copies) / Poly(ADP-ribose) polymerase catalytic domain / Poly(ADP-ribose) polymerase, catalytic domain / PARP catalytic domain profile. / SAM domain (Sterile alpha motif) / SAM domain profile. / Sterile alpha motif. / Sterile alpha motif domain / Sterile alpha motif/pointed domain superfamily ...Ankyrin repeat / Ankyrin repeats (many copies) / Poly(ADP-ribose) polymerase catalytic domain / Poly(ADP-ribose) polymerase, catalytic domain / PARP catalytic domain profile. / SAM domain (Sterile alpha motif) / SAM domain profile. / Sterile alpha motif. / Sterile alpha motif domain / Sterile alpha motif/pointed domain superfamily / Ankyrin repeat / Ankyrin repeats (3 copies) / Ankyrin repeat profile. / Ankyrin repeat region circular profile. / ankyrin repeats / Ankyrin repeat / Ankyrin repeat-containing domain superfamily
Similarity search - Domain/homology
Poly [ADP-ribose] polymerase tankyrase-2
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodhelical reconstruction / cryo EM / Resolution: 2.98 Å
AuthorsMariotti L / Inian O / Desfosses A / Beuron F / Morris EP / Guettler S
Funding support United Kingdom, 4 items
OrganizationGrant numberCountry
Cancer Research UKC47521/A28286 United Kingdom
Wellcome Trust214311/Z/18/Z United Kingdom
The Institute of Cancer Research (ICR)n.a. United Kingdom
The Lister Institute of Preventive Medicinen.a. United Kingdom
CitationJournal: Nature / Year: 2022
Title: Structural basis of tankyrase activation by polymerization.
Authors: Nisha Pillay / Laura Mariotti / Mariola Zaleska / Oviya Inian / Matthew Jessop / Sam Hibbs / Ambroise Desfosses / Paul C R Hopkins / Catherine M Templeton / Fabienne Beuron / Edward P Morris ...Authors: Nisha Pillay / Laura Mariotti / Mariola Zaleska / Oviya Inian / Matthew Jessop / Sam Hibbs / Ambroise Desfosses / Paul C R Hopkins / Catherine M Templeton / Fabienne Beuron / Edward P Morris / Sebastian Guettler /
Abstract: The poly-ADP-ribosyltransferase tankyrase (TNKS, TNKS2) controls a wide range of disease-relevant cellular processes, including WNT-β-catenin signalling, telomere length maintenance, Hippo ...The poly-ADP-ribosyltransferase tankyrase (TNKS, TNKS2) controls a wide range of disease-relevant cellular processes, including WNT-β-catenin signalling, telomere length maintenance, Hippo signalling, DNA damage repair and glucose homeostasis. This has incentivized the development of tankyrase inhibitors. Notwithstanding, our knowledge of the mechanisms that control tankyrase activity has remained limited. Both catalytic and non-catalytic functions of tankyrase depend on its filamentous polymerization. Here we report the cryo-electron microscopy reconstruction of a filament formed by a minimal active unit of tankyrase, comprising the polymerizing sterile alpha motif (SAM) domain and its adjacent catalytic domain. The SAM domain forms a novel antiparallel double helix, positioning the protruding catalytic domains for recurring head-to-head and tail-to-tail interactions. The head interactions are highly conserved among tankyrases and induce an allosteric switch in the active site within the catalytic domain to promote catalysis. Although the tail interactions have a limited effect on catalysis, they are essential to tankyrase function in WNT-β-catenin signalling. This work reveals a novel SAM domain polymerization mode, illustrates how supramolecular assembly controls catalytic and non-catalytic functions, provides important structural insights into the regulation of a non-DNA-dependent poly-ADP-ribosyltransferase and will guide future efforts to modulate tankyrase and decipher its contribution to disease mechanisms.
History
DepositionAug 1, 2022-
Header (metadata) releaseNov 16, 2022-
Map releaseNov 16, 2022-
UpdateDec 14, 2022-
Current statusDec 14, 2022Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_15520.png

Downloads & links

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Map

FileDownload / File: emd_15520.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesX (Sec.)Y (Row.)Z (Col.)
1.06 Å/pix.
x 400 pix.
= 424. Å		1.06 Å/pix.
x 400 pix.
= 424. Å		1.06 Å/pix.
x 400 pix.
= 424. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.06 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 8.73
Minimum - Maximum-22.093016 - 41.711735
Average (Standard dev.)-0.0006398231 (±0.9761947)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderZYX
Origin000
Dimensions400400400
Spacing400400400
CellA=B=C: 423.99997 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : TNKS2 SAM-PARP (867-1162) filament

EntireName: TNKS2 SAM-PARP (867-1162) filament
Components
  • Complex: TNKS2 SAM-PARP (867-1162) filament
    • Protein or peptide: Poly [ADP-ribose] polymerase tankyrase-2
  • Ligand: ZINC ION

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Supramolecule #1: TNKS2 SAM-PARP (867-1162) filament

SupramoleculeName: TNKS2 SAM-PARP (867-1162) filament / type: complex / Chimera: Yes / ID: 1 / Parent: 0 / Macromolecule list: #1
Details: double-helical filament of human TNKS2 SAM-PARP G1032W, residues 867-1162, N-terminal vector-derived SNA tripeptide, 20 protomers in refined structure
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 34 kDa/nm

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Macromolecule #1: Poly [ADP-ribose] polymerase tankyrase-2

MacromoleculeName: Poly [ADP-ribose] polymerase tankyrase-2 / type: protein_or_peptide / ID: 1 / Number of copies: 20 / Enantiomer: LEVO / EC number: NAD+ ADP-ribosyltransferase
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 34.077668 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: SNAEKKEVPG VDFSITQFVR NLGLEHLMDI FEREQITLDV LVEMGHKELK EIGINAYGHR HKLIKGVERL ISGQQGLNPY LTLNTSGSG TILIDLSPDD KEFQSVEEEM QSTVREHRDG GHAGGIFNRY NILKIQKVCN KKLWERYTHR RKEVSEENHN H ANERMLFH ...String:
SNAEKKEVPG VDFSITQFVR NLGLEHLMDI FEREQITLDV LVEMGHKELK EIGINAYGHR HKLIKGVERL ISGQQGLNPY LTLNTSGSG TILIDLSPDD KEFQSVEEEM QSTVREHRDG GHAGGIFNRY NILKIQKVCN KKLWERYTHR RKEVSEENHN H ANERMLFH WSPFVNAIIH KGFDERHAYI GGMFGAGIYF AENSSKSNQY VYGIGGGTGC PVHKDRSCYI CHRQLLFCRV TL GKSFLQF SAMKMAHSPP GHHSVTGRPS VNGLALAEYV IYRGEQAYPE YLITYQIMRP EG

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Macromolecule #2: ZINC ION

MacromoleculeName: ZINC ION / type: ligand / ID: 2 / Number of copies: 20 / Formula: ZN
Molecular weightTheoretical: 65.409 Da

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Experimental details

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Structure determination

Methodcryo EM
Processinghelical reconstruction
Aggregation statefilament

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Sample preparation

Concentration0.86 mg/mL
BufferpH: 7.5
Component:
ConcentrationFormulaName
50.0 mMTris-HClTrisTris buffer
500.0 mMNaClSodium chloridesodium chloride
10.0 mMbeta-MEbeta-mercaptoethanol2-Mercaptoethanol

Details: After brief incubation on the grid, the sample was washed 10 times with water to gradually lower the salt concentration and improve sample contrast.
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Instrument: FEI VITROBOT MARK IV
DetailsThe PARP domain is inactivated by a G1032W mutation.

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Electron microscopy

MicroscopeTFS KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 3.5 µm / Nominal defocus min: 1.2 µm / Nominal magnification: 81000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
TemperatureMin: 80.0 K / Max: 80.0 K
Image recording#0 - Image recording ID: 1 / #0 - Film or detector model: GATAN K2 SUMMIT (4k x 4k) / #0 - Detector mode: INTEGRATING / #0 - Average electron dose: 40.0 e/Å2 / #1 - Image recording ID: 2 / #1 - Film or detector model: GATAN K3 (6k x 4k) / #1 - Average electron dose: 40.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Segment selectionNumber selected: 188925
Final angle assignmentType: NOT APPLICABLE
Final reconstructionApplied symmetry - Helical parameters - Δz: 13.6 Å
Applied symmetry - Helical parameters - Δ&Phi: -52.3 °
Applied symmetry - Helical parameters - Axial symmetry: D1 (2x1 fold dihedral)
Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 2.98 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.08) / Number images used: 139880
Image recording ID2
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial model
PDB IDChain

5nwg

chain_id: IB

5jrt

chain_id: A
RefinementSpace: REAL / Protocol: FLEXIBLE FIT / Overall B value: 7.41 / Target criteria: FSC
Output model

PDB-8aly:
Cryo-EM structure of human tankyrase 2 SAM-PARP filament (G1032W mutant)

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