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- EMDB-9931: Cryo-EM structure of the human P4-type flippase ATP8A1-CDC50 (E1 ... -

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Basic information

Entry
Database: EMDB / ID: EMD-9931
TitleCryo-EM structure of the human P4-type flippase ATP8A1-CDC50 (E1 state class1)
Map data
SampleATP8A1-CDC50a:
Phospholipid-transporting ATPase / Cell cycle control protein 50A / (ligand) x 3
Function / homology
Function and homology information


ATPase-coupled intramembrane lipid transporter activity / aminophospholipid flippase activity / aminophospholipid transport / positive regulation of protein exit from endoplasmic reticulum / protein localization to endosome / P-type phospholipid transporter / phospholipid transport / phospholipid translocation / azurophil granule membrane / drug transmembrane transport ...ATPase-coupled intramembrane lipid transporter activity / aminophospholipid flippase activity / aminophospholipid transport / positive regulation of protein exit from endoplasmic reticulum / protein localization to endosome / P-type phospholipid transporter / phospholipid transport / phospholipid translocation / azurophil granule membrane / drug transmembrane transport / transport vesicle membrane / specific granule membrane / positive regulation of neuron projection development / apical plasma membrane / neutrophil degranulation / endoplasmic reticulum / Golgi apparatus / magnesium ion binding / membrane / integral component of membrane / ATP binding / plasma membrane
P-type ATPase, A domain superfamily / CDC50/LEM3 family / P-type ATPase, phosphorylation site / P-type ATPase, transmembrane domain superfamily / P-type ATPase, cytoplasmic domain N / P-type ATPase / HAD superfamily / P-type ATPase, subfamily IV / Cell cycle control protein 50A / P-type ATPase, C-terminal ...P-type ATPase, A domain superfamily / CDC50/LEM3 family / P-type ATPase, phosphorylation site / P-type ATPase, transmembrane domain superfamily / P-type ATPase, cytoplasmic domain N / P-type ATPase / HAD superfamily / P-type ATPase, subfamily IV / Cell cycle control protein 50A / P-type ATPase, C-terminal / P-type ATPase, N-terminal / HAD-like superfamily
Phospholipid-transporting ATPase / Cell cycle control protein 50A
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsHiraizumi M / Yamashita K / Nishizawa T / Nureki O
CitationJournal: Science / Year: 2019
Title: Cryo-EM structures capture the transport cycle of the P4-ATPase flippase.
Authors: Masahiro Hiraizumi / Keitaro Yamashita / Tomohiro Nishizawa / Osamu Nureki /
Abstract: In eukaryotic membranes, type IV P-type adenosine triphosphatases (P4-ATPases) mediate the translocation of phospholipids from the outer to the inner leaflet and maintain lipid asymmetry, which is ...In eukaryotic membranes, type IV P-type adenosine triphosphatases (P4-ATPases) mediate the translocation of phospholipids from the outer to the inner leaflet and maintain lipid asymmetry, which is critical for membrane trafficking and signaling pathways. Here, we report the cryo-electron microscopy structures of six distinct intermediates of the human ATP8A1-CDC50a heterocomplex at resolutions of 2.6 to 3.3 angstroms, elucidating the lipid translocation cycle of this P4-ATPase. ATP-dependent phosphorylation induces a large rotational movement of the actuator domain around the phosphorylation site in the phosphorylation domain, accompanied by lateral shifts of the first and second transmembrane helices, thereby allowing phosphatidylserine binding. The phospholipid head group passes through the hydrophilic cleft, while the acyl chain is exposed toward the lipid environment. These findings advance our understanding of the flippase mechanism and the disease-associated mutants of P4-ATPases.
Validation ReportPDB-ID: 6k7g

SummaryFull reportAbout validation report
History
DepositionJun 7, 2019-
Header (metadata) releaseAug 21, 2019-
Map releaseAug 28, 2019-
UpdateNov 13, 2019-
Current statusNov 13, 2019Processing site: PDBj / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.015
  • Imaged by UCSF Chimera
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  • Surface view colored by height
  • Surface level: 0.015
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: : PDB-6k7g
  • Surface level: 0.015
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_9931.png

Downloads & links

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Map

FileDownload / File: emd_9931.map.gz / Format: CCP4 / Size: 67 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.83 Å/pix.
x 260 pix.
= 215.8 Å		0.83 Å/pix.
x 260 pix.
= 215.8 Å		0.83 Å/pix.
x 260 pix.
= 215.8 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.83 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.015 / Movie #1: 0.015
Minimum - Maximum-0.08463873 - 0.14112954
Average (Standard dev.)0.00026575278 (±0.003903605)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions260260260
Spacing260260260
CellA=B=C: 215.8 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.830.830.83
M x/y/z260260260
origin x/y/z0.0000.0000.000
length x/y/z215.800215.800215.800
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ200200200
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS260260260
D min/max/mean-0.0850.1410.000

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Supplemental data

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Sample components

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Entire ATP8A1-CDC50a

EntireName: ATP8A1-CDC50a / Number of components: 6

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Component #1: protein, ATP8A1-CDC50a

ProteinName: ATP8A1-CDC50a / Recombinant expression: No
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Homo sapiens (human) / Vector: pcDNA3.4 / Cell of expression system: Expi293F

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Component #2: protein, Phospholipid-transporting ATPase

ProteinName: Phospholipid-transporting ATPase / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 130.537852 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Homo sapiens (human)

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Component #3: protein, Cell cycle control protein 50A

ProteinName: Cell cycle control protein 50A / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 40.727527 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Homo sapiens (human)

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Component #4: ligand, N-ACETYL-D-GLUCOSAMINE

LigandName: N-ACETYL-D-GLUCOSAMINEN-Acetylglucosamine / Number of Copies: 4 / Recombinant expression: No
MassTheoretical: 0.221208 kDa

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Component #5: ligand, ALPHA-D-MANNOSE

LigandName: ALPHA-D-MANNOSE / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 0.180156 kDa

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Component #6: ligand, CHOLESTEROL HEMISUCCINATE

LigandName: CHOLESTEROL HEMISUCCINATE / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 0.486726 kDa

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Experimental details

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Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionpH: 7.5
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 64 e/Å2 / Illumination mode: OTHER
LensImaging mode: BRIGHT FIELD
Specimen HolderModel: OTHER
CameraDetector: OTHER

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Image processing

ProcessingMethod: single particle reconstruction / Number of projections: 139434
3D reconstructionResolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF
FSC plot (resolution estimation)

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Atomic model buiding

Output model

6k7g

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