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- PDB-9zpd: Cryo-EM structure of human PI3KC3-C2 in complex with Rubicon Midd... -

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Basic information

Entry
Database: PDB / ID: 9zpd
TitleCryo-EM structure of human PI3KC3-C2 in complex with Rubicon Middle Region of C terminus
Components
  • Beclin-1,Run domain Beclin-1-interacting and cysteine-rich domain-containing protein chimera
  • Phosphatidylinositol 3-kinase catalytic subunit type 3
  • Phosphoinositide 3-kinase regulatory subunit 4
  • UV radiation resistance-associated gene protein
KeywordsSIGNALING PROTEIN / Lipid kinase / endocytic sorting / cytokinesis / autophagosome maturation / lysosome recycling / LC3-associated phagocytosis
Function / homology
Function and homology information


negative regulation of autophagosome maturation / regulation of protein serine/threonine kinase activity / lytic vacuole / maintenance of Golgi location / phosphatidylinositol 3-kinase inhibitor activity / cellular response to aluminum ion / positive regulation of protein lipidation / postsynaptic endosome / Toll Like Receptor 9 (TLR9) Cascade / Synthesis of PIPs at the late endosome membrane ...negative regulation of autophagosome maturation / regulation of protein serine/threonine kinase activity / lytic vacuole / maintenance of Golgi location / phosphatidylinositol 3-kinase inhibitor activity / cellular response to aluminum ion / positive regulation of protein lipidation / postsynaptic endosome / Toll Like Receptor 9 (TLR9) Cascade / Synthesis of PIPs at the late endosome membrane / positive regulation of stress granule assembly / phosphatidylinositol 3-kinase complex, class III / cellular response to oxygen-glucose deprivation / Synthesis of PIPs at the early endosome membrane / phosphatidylinositol 3-kinase complex, class III, type II / phosphatidylinositol 3-kinase complex, class III, type I / phosphatidylinositol phosphate binding / response to mitochondrial depolarisation / SARS-CoV-2 modulates autophagy / host-mediated activation of viral genome replication / presynaptic endosome / positive regulation of attachment of mitotic spindle microtubules to kinetochore / negative regulation of lysosome organization / engulfment of apoptotic cell / Synthesis of PIPs at the Golgi membrane / phosphatidylinositol kinase activity / positive regulation of autophagosome assembly / response to L-leucine / early endosome to late endosome transport / cytoplasmic side of mitochondrial outer membrane / negative regulation of autophagosome assembly / protein localization to phagophore assembly site / multivesicular body sorting pathway / receptor catabolic process / SMAD protein signal transduction / protein targeting to lysosome / late endosome to vacuole transport / negative regulation of endocytosis / endosome organization / pexophagy / double-strand break repair via classical nonhomologous end joining / retrograde vesicle-mediated transport, Golgi to endoplasmic reticulum / phagophore assembly site / Translation of Replicase and Assembly of the Replication Transcription Complex / positive regulation of autophagosome maturation / phosphatidylinositol-3-phosphate biosynthetic process / cellular response to nitrogen starvation / centrosome cycle / negative regulation of programmed cell death / phosphatidylinositol 3-kinase / spindle organization / 1-phosphatidylinositol-3-kinase activity / lysosome organization / response to vitamin E / mitotic metaphase chromosome alignment / cytoplasmic pattern recognition receptor signaling pathway / Macroautophagy / phosphatidylinositol-mediated signaling / response to iron(II) ion / positive regulation of cardiac muscle hypertrophy / SNARE complex assembly / RSV-host interactions / p38MAPK cascade / phosphatidylinositol phosphate biosynthetic process / autolysosome / autophagosome membrane / PI3K Cascade / axoneme / chromosome, centromeric region / amyloid-beta metabolic process / autophagosome maturation / autophagosome assembly / RHO GTPases Activate NADPH Oxidases / immune system process / synaptic vesicle endocytosis / regulation of macroautophagy / phagocytosis / neuron development / cellular defense response / phosphatidylinositol 3-kinase binding / cellular response to glucose starvation / mitophagy / positive regulation of intrinsic apoptotic signaling pathway / phagocytic vesicle / JNK cascade / endoplasmic reticulum-Golgi intermediate compartment membrane / positive regulation of autophagy / negative regulation of phosphatidylinositol 3-kinase/protein kinase B signal transduction / negative regulation of autophagy / autophagosome / cellular response to amino acid starvation / cellular response to copper ion / cellular response to epidermal growth factor stimulus / SNARE binding / regulation of cytokinesis / Antigen Presentation: Folding, assembly and peptide loading of class I MHC / macroautophagy / chromosome segregation / phosphatidylinositol 3-kinase/protein kinase B signal transduction / trans-Golgi network
Similarity search - Function
: / : / Rubicon, PI3K-binding domain / Rubicon Homology Domain / Rubicon Homology Domain / DUF4206 / UV radiation resistance protein/autophagy-related protein 14 / Vacuolar sorting 38 and autophagy-related subunit 14 / Serine/threonine-protein kinase Vps15-like / : ...: / : / Rubicon, PI3K-binding domain / Rubicon Homology Domain / Rubicon Homology Domain / DUF4206 / UV radiation resistance protein/autophagy-related protein 14 / Vacuolar sorting 38 and autophagy-related subunit 14 / Serine/threonine-protein kinase Vps15-like / : / VPS15-like, helical domain / RUN / RUN domain / RUN domain superfamily / RUN domain / RUN domain profile. / Beclin-1, BH3 domain / Beclin-1 BH3 domain, Bcl-2-interacting / Atg6/Beclin / Atg6/Beclin C-terminal domain superfamily / Atg6, BARA domain / Atg6/beclin, coiled-coil domain / Apg6 BARA domain / Apg6 coiled-coil region / Phosphatidylinositol 3-kinase, Vps34 type / HEAT repeat profile. / HEAT, type 2 / C2 phosphatidylinositol 3-kinase-type domain / Phosphoinositide 3-kinase C2 / C2 phosphatidylinositol 3-kinase (PI3K)-type domain profile. / Phosphoinositide 3-kinase, region postulated to contain C2 domain / Phosphoinositide 3-kinase family, accessory domain (PIK domain) / Phosphoinositide 3-kinase family, accessory domain (PIK domain) / Phosphoinositide 3-kinase, accessory (PIK) domain superfamily / Phosphoinositide 3-kinase, accessory (PIK) domain / Phosphatidylinositol kinase / PIK helical domain profile. / Protein kinase C conserved region 2 (CalB) / C2 domain / C2 domain / C2 domain profile. / Phosphatidylinositol 3- and 4-kinases signature 1. / Phosphatidylinositol 3/4-kinase, conserved site / Phosphatidylinositol 3- and 4-kinases signature 2. / Phosphatidylinositol 3-/4-kinase, catalytic domain superfamily / Phosphoinositide 3-kinase, catalytic domain / Phosphatidylinositol 3- and 4-kinase / Phosphatidylinositol 3- and 4-kinases catalytic domain profile. / Phosphatidylinositol 3-/4-kinase, catalytic domain / C2 domain superfamily / Armadillo-like helical / WD domain, G-beta repeat / Armadillo-type fold / Serine/threonine-protein kinase, active site / Serine/Threonine protein kinases active-site signature. / Trp-Asp (WD) repeats profile. / Trp-Asp (WD) repeats circular profile. / WD40 repeats / WD40 repeat / Protein kinase domain / WD40-repeat-containing domain superfamily / Serine/Threonine protein kinases, catalytic domain / WD40/YVTN repeat-like-containing domain superfamily / Protein kinase domain profile. / Protein kinase domain / Protein kinase-like domain superfamily
Similarity search - Domain/homology
GUANOSINE-5'-TRIPHOSPHATE / Phosphoinositide 3-kinase regulatory subunit 4 / Beclin-1 / Phosphatidylinositol 3-kinase catalytic subunit type 3 / Run domain Beclin-1-interacting and cysteine-rich domain-containing protein / UV radiation resistance-associated gene protein
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.38 Å
AuthorsChen, M. / Ni, Q. / Hurley, J.H.
Funding support United States, 2items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)NS134598 United States
Michael J. Fox FoundationASAP-000350 United States
CitationJournal: To Be Published
Title: Cryo-EM reconstruction of PI3KC3-C2 in complex with Rubicon Middle Region of C terminus
Authors: Chen, M. / Ni, Q. / Hurley, J.H.
History
DepositionDec 16, 2025Deposition site: RCSB / Processing site: RCSB
Revision 1.0Dec 24, 2025Provider: repository / Type: Initial release
Revision 1.0Dec 24, 2025Data content type: EM metadata / Data content type: EM metadata / Provider: repository / Type: Initial release
Revision 1.0Dec 24, 2025Data content type: FSC / Data content type: FSC / Provider: repository / Type: Initial release
Revision 1.0Dec 24, 2025Data content type: Half map / Part number: 1 / Data content type: Half map / Provider: repository / Type: Initial release
Revision 1.0Dec 24, 2025Data content type: Half map / Part number: 2 / Data content type: Half map / Provider: repository / Type: Initial release
Revision 1.0Dec 24, 2025Data content type: Image / Data content type: Image / Provider: repository / Type: Initial release
Revision 1.0Dec 24, 2025Data content type: Primary map / Data content type: Primary map / Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Phosphoinositide 3-kinase regulatory subunit 4
B: Phosphatidylinositol 3-kinase catalytic subunit type 3
C: UV radiation resistance-associated gene protein
D: Beclin-1,Run domain Beclin-1-interacting and cysteine-rich domain-containing protein chimera
E: Beclin-1,Run domain Beclin-1-interacting and cysteine-rich domain-containing protein chimera
hetero molecules


Theoretical massNumber of molelcules
Total (without water)493,9677
Polymers493,3795
Non-polymers5892
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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Protein , 4 types, 5 molecules ABCDE

#1: Protein Phosphoinositide 3-kinase regulatory subunit 4


Mass: 162838.141 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: PIK3R4 / Production host: Homo sapiens (human)
References: UniProt: A0JP11, non-specific serine/threonine protein kinase
#2: Protein Phosphatidylinositol 3-kinase catalytic subunit type 3 / PI3-kinase type 3 / PI3K type 3 / PtdIns-3-kinase type 3 / Phosphatidylinositol 3-kinase p100 ...PI3-kinase type 3 / PI3K type 3 / PtdIns-3-kinase type 3 / Phosphatidylinositol 3-kinase p100 subunit / Phosphoinositide-3-kinase class 3 / hVps34


Mass: 101680.328 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: PIK3C3, VPS34 / Production host: Homo sapiens (human) / References: UniProt: Q8NEB9, phosphatidylinositol 3-kinase
#3: Protein UV radiation resistance-associated gene protein / p63


Mass: 78258.836 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: UVRAG / Production host: Homo sapiens (human) / References: UniProt: Q9P2Y5
#4: Protein Beclin-1,Run domain Beclin-1-interacting and cysteine-rich domain-containing protein chimera / Coiled-coil myosin-like BCL2-interacting protein / Protein GT197 / Rubicon / Beclin-1 associated ...Coiled-coil myosin-like BCL2-interacting protein / Protein GT197 / Rubicon / Beclin-1 associated RUN domain containing protein / Baron


Mass: 75300.797 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: BECN1, GT197, RUBCN, KIAA0226 / Production host: Homo sapiens (human) / References: UniProt: Q14457, UniProt: Q92622

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Non-polymers , 2 types, 2 molecules

#5: Chemical ChemComp-GTP / GUANOSINE-5'-TRIPHOSPHATE


Mass: 523.180 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C10H16N5O14P3 / Feature type: SUBJECT OF INVESTIGATION / Comment: GTP, energy-carrying molecule*YM
#6: Chemical ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Zn / Feature type: SUBJECT OF INVESTIGATION

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Details

Has ligand of interestY
Has protein modificationN

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Experimental details

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Experiment

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

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Sample preparation

ComponentName: Human Phosphatidylinositol-3 kinase class III complex II in complex with Rubicon Middle Region of C terminus
Type: COMPLEX / Entity ID: #1-#4 / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Homo sapiens (human)
Buffer solutionpH: 7.5
Buffer component
IDConc.NameFormulaBuffer-ID
1200 mMSodium chlorideNaCl1
22 mMMagnesium chlorideMgCl21
325 mM4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid1
41 mMTris(2-carboxyethyl)phosphine1
SpecimenConc.: 0.15 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 200 divisions/in. / Grid type: Quantifoil Active R2/1
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2000 nm / Nominal defocus min: 800 nm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

EM software
IDNameVersionCategory
1Topaz0.2.5aparticle selection
2SerialEM4.0.20image acquisition
4cryoSPARC4.5.3CTF correction
7Coot0.9.8model fitting
9PHENIX1.21.2_5419model refinement
10cryoSPARC4.5.3initial Euler assignment
11cryoSPARC4.5.3final Euler assignment
12cryoSPARC4.5.3classification
13cryoSPARC4.5.33D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.38 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 175726 / Symmetry type: POINT
Atomic model buildingB value: 95.23 / Protocol: FLEXIBLE FIT / Space: REAL
Atomic model buildingSource name: AlphaFold / Type: in silico model

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