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- EMDB-42807: Structure of ADP-bound and phosphorylated Pediculus humanus (Ph) ... -

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Basic information

Entry
Database: EMDB / ID: EMD-42807
TitleStructure of ADP-bound and phosphorylated Pediculus humanus (Ph) PINK1 dimer
Map dataMap (sharpened) of the ADP-bound phosphorylated PhPINK1 dimer
Sample
  • Complex: ADP-bound and phosphorylated Pediculus humanus (Ph) PINK1 dodecamer
    • Protein or peptide: Serine/threonine-protein kinase Pink1, mitochondrial
  • Ligand: ADENOSINE-5'-DIPHOSPHATE
  • Ligand: MAGNESIUM ION
KeywordsPINK1 / Kinase / Mitophagy / Parkinson's Disease / Ubiquitin / Phosphorylation / Phospho-ubiquitin / TRANSFERASE
Function / homology
Function and homology information


autophagy of mitochondrion / positive regulation of mitochondrial fission / regulation of apoptotic process / mitochondrial outer membrane / mitochondrial inner membrane / non-specific serine/threonine protein kinase / protein kinase activity / protein serine/threonine kinase activity / ATP binding / metal ion binding / cytosol
Similarity search - Function
: / Serine/threonine-protein kinase, active site / Serine/Threonine protein kinases active-site signature. / Protein kinase domain / Serine/Threonine protein kinases, catalytic domain / Protein kinase domain profile. / Protein kinase domain / Protein kinase-like domain superfamily
Similarity search - Domain/homology
Serine/threonine-protein kinase Pink1, mitochondrial
Similarity search - Component
Biological speciesPediculus humanus corporis (human body louse)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.13 Å
AuthorsGan ZY / Kirk NS / Leis A / Komander D
Funding support Australia, United States, 2 items
OrganizationGrant numberCountry
National Health and Medical Research Council (NHMRC, Australia) Australia
Michael J. Fox Foundation United States
CitationJournal: Sci Adv / Year: 2024
Title: Interaction of PINK1 with nucleotides and kinetin.
Authors: Zhong Yan Gan / Sylvie Callegari / Thanh N Nguyen / Nicholas S Kirk / Andrew Leis / Michael Lazarou / Grant Dewson / David Komander /
Abstract: The ubiquitin kinase PINK1 accumulates on damaged mitochondria to trigger mitophagy, and PINK1 loss-of-function mutations cause early onset Parkinson's disease. Nucleotide analogs such as kinetin ...The ubiquitin kinase PINK1 accumulates on damaged mitochondria to trigger mitophagy, and PINK1 loss-of-function mutations cause early onset Parkinson's disease. Nucleotide analogs such as kinetin triphosphate (KTP) were reported to enhance PINK1 activity and may represent a therapeutic strategy for the treatment of Parkinson's disease. Here, we investigate the interaction of PINK1 with nucleotides, including KTP. We establish a cryo-EM platform exploiting the dodecamer assembly of () PINK1 and determine PINK1 structures bound to AMP-PNP and ADP, revealing conformational changes in the kinase N-lobe that help establish PINK1's ubiquitin binding site. Notably, we find that KTP is unable to bind PINK1 or human () PINK1 due to a steric clash with the kinase "gatekeeper" methionine residue, and mutation to Ala or Gly is required for PINK1 to bind and use KTP as a phosphate donor in ubiquitin phosphorylation and mitophagy. PINK1 M318G can be used to conditionally uncouple PINK1 stabilization and activity on mitochondria.
History
DepositionNov 13, 2023-
Header (metadata) releaseJan 31, 2024-
Map releaseJan 31, 2024-
UpdateNov 13, 2024-
Current statusNov 13, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_42807.png

Downloads & links

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Map

FileDownload / File: emd_42807.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationMap (sharpened) of the ADP-bound phosphorylated PhPINK1 dimer
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.81 Å/pix.
x 256 pix.
= 206.848 Å		0.81 Å/pix.
x 256 pix.
= 206.848 Å		0.81 Å/pix.
x 256 pix.
= 206.848 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.808 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.12
Minimum - Maximum-0.47722015 - 0.62876177
Average (Standard dev.)0.0013259078 (±0.017578008)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 206.848 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: Half map of the ADP-bound phosphorylated PhPINK1 dimer

Fileemd_42807_half_map_1.map
AnnotationHalf map of the ADP-bound phosphorylated PhPINK1 dimer
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map of the ADP-bound phosphorylated PhPINK1 dimer

Fileemd_42807_half_map_2.map
AnnotationHalf map of the ADP-bound phosphorylated PhPINK1 dimer
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : ADP-bound and phosphorylated Pediculus humanus (Ph) PINK1 dodecamer

EntireName: ADP-bound and phosphorylated Pediculus humanus (Ph) PINK1 dodecamer
Components
  • Complex: ADP-bound and phosphorylated Pediculus humanus (Ph) PINK1 dodecamer
    • Protein or peptide: Serine/threonine-protein kinase Pink1, mitochondrial
  • Ligand: ADENOSINE-5'-DIPHOSPHATE
  • Ligand: MAGNESIUM ION

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Supramolecule #1: ADP-bound and phosphorylated Pediculus humanus (Ph) PINK1 dodecamer

SupramoleculeName: ADP-bound and phosphorylated Pediculus humanus (Ph) PINK1 dodecamer
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Pediculus humanus corporis (human body louse)

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Macromolecule #1: Serine/threonine-protein kinase Pink1, mitochondrial

MacromoleculeName: Serine/threonine-protein kinase Pink1, mitochondrial / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Pediculus humanus corporis (human body louse)
Molecular weightTheoretical: 53.133582 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: GPSGLLTKDD ELEGICWEIR EAVSKGKWND SESENVEQLQ AANLDELDLG EPIAKGCNAV VYSAKLKNVQ SNKLAHQLAV KMMFNYDVE (SEP)NSTAILKAM YRETVPAMSY FFNQNLFNIE NISDFKIRLP PHPNIVRMYS VFADRIPDLQ CNKQLYP EA LPPRINPEGS ...String:
GPSGLLTKDD ELEGICWEIR EAVSKGKWND SESENVEQLQ AANLDELDLG EPIAKGCNAV VYSAKLKNVQ SNKLAHQLAV KMMFNYDVE (SEP)NSTAILKAM YRETVPAMSY FFNQNLFNIE NISDFKIRLP PHPNIVRMYS VFADRIPDLQ CNKQLYP EA LPPRINPEGS GRNMSLFLVM KRYDCTLKEY LRDK(TPO)PNMRS SILLLSQLLE AVAHMNIHNI SHRDLKSDNI LVDL SEGDA YPTIVITDFG CCLCDKQNGL VIPYRSEDQD KGGNRALMAP EIANAKPGTF SWLNYKKSDL WAVGAIAYEI FNIDN PFYD KTMKLLSKSY KEEDLPELPD TIPFIIRNLV SNMLSRSTNK RLDCDVAATV AQLYLWAPSS WLKENYTLPN SNEIIQ WLL CLSSKVLCER DITARNKTNT MSESVSKAQY KGRRSLPEYE LIASFLRRVR LHLVRKGLKW IQELHIYN

UniProtKB: Serine/threonine-protein kinase Pink1, mitochondrial

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Macromolecule #2: ADENOSINE-5'-DIPHOSPHATE

MacromoleculeName: ADENOSINE-5'-DIPHOSPHATE / type: ligand / ID: 2 / Number of copies: 2 / Formula: ADP
Molecular weightTheoretical: 427.201 Da
Chemical component information

ChemComp-ADP:
ADENOSINE-5'-DIPHOSPHATE / ADP, energy-carrying molecule*YM

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Macromolecule #3: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 3 / Number of copies: 4 / 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

Concentration1.9 mg/mL
BufferpH: 8.5
Component:
ConcentrationNameFormula
25.0 mMtrisaminomethane
150.0 mMsodium chlorideNaCl
10.0 mMdithiothreitol
10.0 mMmagnesium chlorideMgCl2
10.0 mMadenosine diphosphate
GridModel: UltrAuFoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: GOLD / Support film - topology: HOLEY ARRAY / Pretreatment - Time: 120 sec. / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Slit width: 20 eV
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: OTHER / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 1.5 µm / Nominal defocus min: 0.5 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.13 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.2.1) / Number images used: 130515
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.2.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.2.1)
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

7t4n


Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementSpace: REAL
Output model

PDB-8uyi:
Structure of ADP-bound and phosphorylated Pediculus humanus (Ph) PINK1 dimer

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