[English] 日本語
Yorodumi
- PDB-7pem: Cryo-EM structure of phophorylated Drs2p-Cdc50p in a PS and ATP-b... -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: PDB / ID: 7pem
TitleCryo-EM structure of phophorylated Drs2p-Cdc50p in a PS and ATP-bound E2P state
Components
  • Cell division control protein 50
  • Probable phospholipid-transporting ATPase DRS2
KeywordsMEMBRANE PROTEIN / Lipid Flippase / P4 ATPase / trans-Golgi Network / Phosphatidylserine transport
Function / homology
Function and homology information


Cdc50p-Drs2p complex / actin cortical patch localization / Ion transport by P-type ATPases / aminophospholipid translocation / phosphatidylcholine flippase activity / post-Golgi vesicle-mediated transport / phosphatidylserine flippase activity / phosphatidylserine floppase activity / phospholipid-translocating ATPase complex / ATPase-coupled intramembrane lipid transporter activity ...Cdc50p-Drs2p complex / actin cortical patch localization / Ion transport by P-type ATPases / aminophospholipid translocation / phosphatidylcholine flippase activity / post-Golgi vesicle-mediated transport / phosphatidylserine flippase activity / phosphatidylserine floppase activity / phospholipid-translocating ATPase complex / ATPase-coupled intramembrane lipid transporter activity / phosphatidylethanolamine flippase activity / endocytic recycling / P-type phospholipid transporter / phosphatidylinositol-4-phosphate binding / retrograde transport, endosome to Golgi / phospholipid translocation / Neutrophil degranulation / intracellular protein transport / trans-Golgi network / endocytosis / late endosome membrane / endosome membrane / Golgi apparatus / magnesium ion binding / endoplasmic reticulum / ATP hydrolysis activity / ATP binding / plasma membrane / cytosol
Similarity search - Function
CDC50/LEM3 family / LEM3 (ligand-effect modulator 3) family / CDC50 family / P-type ATPase, subfamily IV / P-type ATPase, C-terminal / P-type ATPase, N-terminal / Phospholipid-translocating ATPase N-terminal / Phospholipid-translocating P-type ATPase C-terminal / Cation transport ATPase (P-type) / P-type ATPase, haloacid dehalogenase domain / P-type ATPase, phosphorylation site ...CDC50/LEM3 family / LEM3 (ligand-effect modulator 3) family / CDC50 family / P-type ATPase, subfamily IV / P-type ATPase, C-terminal / P-type ATPase, N-terminal / Phospholipid-translocating ATPase N-terminal / Phospholipid-translocating P-type ATPase C-terminal / Cation transport ATPase (P-type) / P-type ATPase, haloacid dehalogenase domain / P-type ATPase, phosphorylation site / P-type ATPase, cytoplasmic domain N / E1-E2 ATPases phosphorylation site. / P-type ATPase, A domain superfamily / P-type ATPase / P-type ATPase, transmembrane domain superfamily / HAD superfamily / HAD-like superfamily
Similarity search - Domain/homology
Chem-2Y5 / ADENOSINE-5'-TRIPHOSPHATE / Chem-Q3G / Phospholipid-transporting ATPase accessory subunit CDC50 / Phospholipid-transporting ATPase DRS2
Similarity search - Component
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.1 Å
AuthorsTimcenko, M. / Wang, Y. / Lyons, J.A. / Nissen, P. / Lindorff-Larsen, K.
Funding support Denmark, 1items
OrganizationGrant numberCountry
LundbeckfondenR155-2015-266 Denmark
CitationJournal: To Be Published
Title: Substrate Transport and Specificity in a Phospholipid Flippase
Authors: Wang, Y. / Lyons, J.A. / Timcenko, M. / Kummerer, F. / de Groot, B.L. / Nissen, P. / Gapsys, V. / Lindorff-Larsen, K.
History
DepositionAug 10, 2021Deposition site: PDBE / Processing site: PDBE
Revision 1.0Aug 24, 2022Provider: repository / Type: Initial release

-
Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

-
Assembly

Deposited unit
A: Probable phospholipid-transporting ATPase DRS2
C: Cell division control protein 50
hetero molecules


Theoretical massNumber of molelcules
Total (without water)202,9049
Polymers199,0442
Non-polymers3,8607
Water362
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area12100 Å2
ΔGint-38 kcal/mol
Surface area61710 Å2
MethodPISA

-
Components

-
Protein , 2 types, 2 molecules AC

#1: Protein Probable phospholipid-transporting ATPase DRS2


Mass: 154006.766 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Details: Drs2p with a Thrombin cleavable C-terminal biotin acceptor domain (BAD) tag, and additional Thrombin cleavage site in the C-terminus.
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Strain: ATCC 204508 / S288c / Gene: DRS2, YAL026C, FUN38 / Plasmid: pYeDP60 / Production host: Saccharomyces cerevisiae (brewer's yeast) / Strain (production host): W303-1A dpep4
References: UniProt: P39524, P-type phospholipid transporter
#2: Protein Cell division control protein 50


Mass: 45037.312 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Details: Cdc50p with Thrombin cleavable C-terminal His-tag
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Strain: ATCC 204508 / S288c / Gene: CDC50, YCR094W, YCR94W / Plasmid: pYeDP60 / Production host: Saccharomyces cerevisiae (brewer's yeast) / Strain (production host): W303-1A dpep4 / References: UniProt: P25656

-
Sugars , 2 types, 3 molecules

#3: Polysaccharide 2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 424.401 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DGlcpNAcb1-4DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/1,2,1/[a2122h-1b_1-5_2*NCC/3=O]/1-1/a4-b1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{}}LINUCSPDB-CARE
#4: Polysaccharide beta-D-mannopyranose-(1-3)-beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4) ...beta-D-mannopyranose-(1-3)-beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 748.682 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DManpb1-3DManpb1-4DGlcpNAcb1-4DGlcpNAcb1-Glycam Condensed SequenceGMML 1.0
WURCS=2.0/2,4,3/[a2122h-1b_1-5_2*NCC/3=O][a1122h-1b_1-5]/1-1-2-2/a4-b1_b4-c1_c3-d1WURCSPDB2Glycan 1.1.0
[]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-Manp]{[(3+1)][b-D-Manp]{}}}}}LINUCSPDB-CARE

-
Non-polymers , 5 types, 6 molecules

#5: Chemical ChemComp-2Y5 / (2R)-1-{[(R)-hydroxy{[(1R,2R,3R,4R,5S,6R)-2,3,5,6-tetrahydroxy-4-(phosphonooxy)cyclohexyl]oxy}phosphoryl]oxy}-3-(octadecanoyloxy)propan-2-yl (5Z,8Z,11Z,14Z)-icosa-5,8,11,14-tetraenoate / Phosphatidylinositol-4-phosphate / PI4P


Mass: 967.108 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C47H84O16P2
#6: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Mg
#7: Chemical ChemComp-ATP / ADENOSINE-5'-TRIPHOSPHATE / Adenosine triphosphate


Mass: 507.181 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C10H16N5O13P3 / Comment: ATP, energy-carrying molecule*YM
#8: Chemical ChemComp-Q3G / O-[(R)-[(2S)-2-(hexadecanoyloxy)-3-(octadecanoyloxy)propoxy](hydroxy)phosphoryl]-D-serine


Mass: 764.022 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C40H78NO10P / Feature type: SUBJECT OF INVESTIGATION / Comment: phospholipid*YM
#9: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Formula: H2O

-
Details

Has ligand of interestY

-
Experimental details

-
Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

-
Sample preparation

ComponentName: Binary complex of Drs2p-Cdc50p with the regulatory lipid PI4P and transport substrate PS
Type: COMPLEX / Entity ID: #1-#2 / Source: RECOMBINANT
Molecular weightValue: 0.18 MDa / Experimental value: NO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast) / Strain: S288c
Source (recombinant)Organism: Saccharomyces cerevisiae (brewer's yeast) / Strain: W303-1A dpep4 / Plasmid: pYeDP60
Buffer solutionpH: 7
Buffer component
IDConc.NameFormulaBuffer-ID
150 mMMOPS-Tris1
2100 mMKCl1
35 mMMgCl21
41 mMDithiothreitolDTT1
50.03 mg/mLlauryl maltose neopentyl glycolLMNG1
SpecimenConc.: 0.6 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: Purified in detergent lauryl maltose neopentyl glycol (LMNG)
Specimen supportGrid material: COPPER / Grid mesh size: 400 divisions/in. / Grid type: C-flat-1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277.15 K
Details: Incubated with 0.1 mg/mL POPS for 1 hour. 3mM ATP was added to the sample just before application to the grid.

-
Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm
Image recordingAverage exposure time: 1.5 sec. / Electron dose: 60 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 9837
EM imaging opticsEnergyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV

-
Processing

Software
NameVersionClassification
phenix.real_space_refine1.18rc7_3834refinement
PHENIX1.18rc7_3834refinement
EM software
IDNameVersionCategoryDetails
1cryoSPARC2particle selection
2EPUimage acquisition
4cryoSPARC2CTF correctionpatch CTF
7PyMOLmodel fitting
8Cootmodel fitting
10cryoSPARC2initial Euler assignmentheterogeneous refinement
11cryoSPARC3final Euler assignmentnon-uniform refinement
12cryoSPARC2classificationheterogeneous refinement
13cryoSPARC33D reconstructionnon-uniform refinement
14PHENIXmodel refinement
CTF correctionType: NONE
Particle selectionNum. of particles selected: 3029402
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 91415 / Details: non-uniform refinement in cryoSPARC v3 / Symmetry type: POINT
Atomic model buildingB value: 69.1 / Space: REAL / Target criteria: correlation coefficient
Details: Molecular dynamics flexible fitting and energy minimization in Gromacs
Atomic model buildingPDB-ID: 6ROJ

6roj

RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 61.24 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.001811883
ELECTRON MICROSCOPYf_angle_d0.441116107
ELECTRON MICROSCOPYf_chiral_restr0.03991836
ELECTRON MICROSCOPYf_plane_restr0.00291999
ELECTRON MICROSCOPYf_dihedral_angle_d22.30461655

+
About Yorodumi

-
News

-
Feb 9, 2022. New format data for meta-information of EMDB entries

New format data for meta-information of EMDB entries

  • Version 3 of the EMDB header file is now the official format.
  • The previous official version 1.9 will be removed from the archive.

Related info.:EMDB header

External links:wwPDB to switch to version 3 of the EMDB data model

-
Aug 12, 2020. Covid-19 info

Covid-19 info

URL: https://pdbj.org/emnavi/covid19.php

New page: Covid-19 featured information page in EM Navigator.

Related info.:Covid-19 info / Mar 5, 2020. Novel coronavirus structure data

+
Mar 5, 2020. Novel coronavirus structure data

Novel coronavirus structure data

Related info.:Yorodumi Speices / Aug 12, 2020. Covid-19 info

External links:COVID-19 featured content - PDBj / Molecule of the Month (242)：Coronavirus Proteases

+
Jan 31, 2019. EMDB accession codes are about to change! (news from PDBe EMDB page)

EMDB accession codes are about to change! (news from PDBe EMDB page)

  • The allocation of 4 digits for EMDB accession codes will soon come to an end. Whilst these codes will remain in use, new EMDB accession codes will include an additional digit and will expand incrementally as the available range of codes is exhausted. The current 4-digit format prefixed with “EMD-” (i.e. EMD-XXXX) will advance to a 5-digit format (i.e. EMD-XXXXX), and so on. It is currently estimated that the 4-digit codes will be depleted around Spring 2019, at which point the 5-digit format will come into force.
  • The EM Navigator/Yorodumi systems omit the EMD- prefix.

Related info.:Q: What is EMD? / ID/Accession-code notation in Yorodumi/EM Navigator

External links:EMDB Accession Codes are Changing Soon! / Contact to PDBj

+
Jul 12, 2017. Major update of PDB

Major update of PDB

  • wwPDB released updated PDB data conforming to the new PDBx/mmCIF dictionary.
  • This is a major update changing the version number from 4 to 5, and with Remediation, in which all the entries are updated.
  • In this update, many items about electron microscopy experimental information are reorganized (e.g. em_software).
  • Now, EM Navigator and Yorodumi are based on the updated data.

External links:wwPDB Remediation / Enriched Model Files Conforming to OneDep Data Standards Now Available in the PDB FTP Archive

-
Yorodumi

Thousand views of thousand structures

  • Yorodumi is a browser for structure data from EMDB, PDB, SASBDB, etc.
  • This page is also the successor to EM Navigator detail page, and also detail information page/front-end page for Omokage search.
  • The word "yorodu" (or yorozu) is an old Japanese word meaning "ten thousand". "mi" (miru) is to see.

Related info.:EMDB / PDB / SASBDB / Comparison of 3 databanks / Yorodumi Search / Aug 31, 2016. New EM Navigator & Yorodumi / Yorodumi Papers / Jmol/JSmol / Function and homology information / Changes in new EM Navigator and Yorodumi

Read more