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- PDB-7x0v: cryo-EM structure of human TRiC-ADP-AlFx -

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Basic information

Entry
Database: PDB / ID: 7x0v
Titlecryo-EM structure of human TRiC-ADP-AlFx
Components(T-complex protein 1 subunit ...) x 8
KeywordsSTRUCTURAL PROTEIN / Human TRiC / tubulin
Function / homology
Function and homology information


zona pellucida receptor complex / scaRNA localization to Cajal body / positive regulation of protein localization to Cajal body / tubulin complex assembly / chaperone mediated protein folding independent of cofactor / positive regulation of establishment of protein localization to telomere / BBSome-mediated cargo-targeting to cilium / chaperonin-containing T-complex / Folding of actin by CCT/TriC / positive regulation of telomerase RNA localization to Cajal body ...zona pellucida receptor complex / scaRNA localization to Cajal body / positive regulation of protein localization to Cajal body / tubulin complex assembly / chaperone mediated protein folding independent of cofactor / positive regulation of establishment of protein localization to telomere / BBSome-mediated cargo-targeting to cilium / chaperonin-containing T-complex / Folding of actin by CCT/TriC / positive regulation of telomerase RNA localization to Cajal body / Formation of tubulin folding intermediates by CCT/TriC / binding of sperm to zona pellucida / Prefoldin mediated transfer of substrate to CCT/TriC / RHOBTB1 GTPase cycle / intermediate filament cytoskeleton / WD40-repeat domain binding / beta-tubulin binding / pericentriolar material / Association of TriC/CCT with target proteins during biosynthesis / chaperone-mediated protein complex assembly / heterochromatin / RHOBTB2 GTPase cycle / chaperone-mediated protein folding / protein folding chaperone / positive regulation of telomerase activity / positive regulation of telomere maintenance via telomerase / Gene and protein expression by JAK-STAT signaling after Interleukin-12 stimulation / acrosomal vesicle / mRNA 3'-UTR binding / cell projection / ATP-dependent protein folding chaperone / response to virus / mRNA 5'-UTR binding / cilium / Cooperation of PDCL (PhLP1) and TRiC/CCT in G-protein beta folding / G-protein beta-subunit binding / azurophil granule lumen / melanosome / unfolded protein binding / protein folding / cell body / secretory granule lumen / microtubule / ficolin-1-rich granule lumen / protein stabilization / cytoskeleton / cadherin binding / centrosome / ubiquitin protein ligase binding / Neutrophil degranulation / Golgi apparatus / ATP hydrolysis activity / RNA binding / extracellular exosome / extracellular region / nucleoplasm / ATP binding / cytosol / cytoplasm
Similarity search - Function
T-complex protein 1, alpha subunit / T-complex protein 1, eta subunit / T-complex protein 1, theta subunit / T-complex protein 1, zeta subunit / T-complex protein 1, delta subunit / T-complex protein 1, gamma subunit / T-complex protein 1, epsilon subunit / T-complex protein 1, beta subunit / Chaperonins TCP-1 signature 1. / Chaperonins TCP-1 signature 2. ...T-complex protein 1, alpha subunit / T-complex protein 1, eta subunit / T-complex protein 1, theta subunit / T-complex protein 1, zeta subunit / T-complex protein 1, delta subunit / T-complex protein 1, gamma subunit / T-complex protein 1, epsilon subunit / T-complex protein 1, beta subunit / Chaperonins TCP-1 signature 1. / Chaperonins TCP-1 signature 2. / Chaperonin TCP-1, conserved site / Chaperonins TCP-1 signature 3. / Chaperone tailless complex polypeptide 1 (TCP-1) / GroEL-like equatorial domain superfamily / TCP-1-like chaperonin intermediate domain superfamily / GroEL-like apical domain superfamily / TCP-1/cpn60 chaperonin family / Chaperonin Cpn60/GroEL/TCP-1 family
Similarity search - Domain/homology
ADENOSINE-5'-DIPHOSPHATE / ALUMINUM FLUORIDE / T-complex protein 1 subunit alpha / T-complex protein 1 subunit zeta / T-complex protein 1 subunit epsilon / T-complex protein 1 subunit gamma / T-complex protein 1 subunit theta / T-complex protein 1 subunit delta / T-complex protein 1 subunit beta / T-complex protein 1 subunit eta
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.2 Å
AuthorsCong, Y. / Liu, C.X.
Funding support China, 1items
OrganizationGrant numberCountry
Chinese Academy of SciencesXDB29040300 China
CitationJournal: Commun Biol / Year: 2023
Title: Pathway and mechanism of tubulin folding mediated by TRiC/CCT along its ATPase cycle revealed using cryo-EM.
Authors: Caixuan Liu / Mingliang Jin / Shutian Wang / Wenyu Han / Qiaoyu Zhao / Yifan Wang / Cong Xu / Lei Diao / Yue Yin / Chao Peng / Lan Bao / Yanxing Wang / Yao Cong /
Abstract: The eukaryotic chaperonin TRiC/CCT assists the folding of about 10% of cytosolic proteins through an ATP-driven conformational cycle, and the essential cytoskeleton protein tubulin is the obligate ...The eukaryotic chaperonin TRiC/CCT assists the folding of about 10% of cytosolic proteins through an ATP-driven conformational cycle, and the essential cytoskeleton protein tubulin is the obligate substrate of TRiC. Here, we present an ensemble of cryo-EM structures of endogenous human TRiC throughout its ATPase cycle, with three of them revealing endogenously engaged tubulin in different folding stages. The open-state TRiC-tubulin-S1 and -S2 maps show extra density corresponding to tubulin in the cis-ring chamber of TRiC. Our structural and XL-MS analyses suggest a gradual upward translocation and stabilization of tubulin within the TRiC chamber accompanying TRiC ring closure. In the closed TRiC-tubulin-S3 map, we capture a near-natively folded tubulin-with the tubulin engaging through its N and C domains mainly with the A and I domains of the CCT3/6/8 subunits through electrostatic and hydrophilic interactions. Moreover, we also show the potential role of TRiC C-terminal tails in substrate stabilization and folding. Our study delineates the pathway and molecular mechanism of TRiC-mediated folding of tubulin along the ATPase cycle of TRiC, and may also inform the design of therapeutic agents targeting TRiC-tubulin interactions.
History
DepositionFeb 22, 2022Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Apr 12, 2023Provider: repository / Type: Initial release
Revision 1.1Jul 5, 2023Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
K: T-complex protein 1 subunit zeta
J: T-complex protein 1 subunit theta
H: T-complex protein 1 subunit eta
G: T-complex protein 1 subunit gamma
E: T-complex protein 1 subunit epsilon
I: T-complex protein 1 subunit delta
B: T-complex protein 1 subunit beta
A: T-complex protein 1 subunit alpha
z: T-complex protein 1 subunit zeta
P: T-complex protein 1 subunit theta
O: T-complex protein 1 subunit eta
N: T-complex protein 1 subunit gamma
e: T-complex protein 1 subunit epsilon
a: T-complex protein 1 subunit alpha
M: T-complex protein 1 subunit delta
L: T-complex protein 1 subunit beta
hetero molecules


Theoretical massNumber of molelcules
Total (without water)955,28464
Polymers946,71616
Non-polymers8,56848
Water28816
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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T-complex protein 1 subunit ... , 8 types, 16 molecules KzJPHOGNEeIMBLAa

#1: Protein T-complex protein 1 subunit zeta / TCP-1-zeta / Acute morphine dependence-related protein 2 / CCT-zeta-1 / HTR3 / Tcp20


Mass: 58106.086 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CCT6A, CCT6, CCTZ / Cell line (production host): HEK293F / Production host: Homo sapiens (human) / References: UniProt: P40227
#2: Protein T-complex protein 1 subunit theta / TCP-1-theta / CCT-theta / Chaperonin containing T-complex polypeptide 1 subunit 8


Mass: 59691.422 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CCT8, C21orf112, CCTQ / Cell line (production host): HEK293F / Production host: Homo sapiens (human) / References: UniProt: P50990
#3: Protein T-complex protein 1 subunit eta / TCP-1-eta / CCT-eta / HIV-1 Nef-interacting protein


Mass: 59417.457 Da / Num. of mol.: 2 / Mutation: L290S
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CCT7, CCTH, NIP7-1 / Cell line (production host): HEK293F / Production host: Homo sapiens (human) / References: UniProt: Q99832
#4: Protein T-complex protein 1 subunit gamma / TCP-1-gamma / CCT-gamma / hTRiC5


Mass: 60613.855 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CCT3, CCTG, TRIC5 / Cell line (production host): HEK293F / Production host: Homo sapiens (human) / References: UniProt: P49368
#5: Protein T-complex protein 1 subunit epsilon / TCP-1-epsilon / CCT-epsilon


Mass: 59547.684 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CCT5, CCTE, KIAA0098 / Cell line (production host): HEK293F / Production host: Homo sapiens (human) / References: UniProt: P48643
#6: Protein T-complex protein 1 subunit delta / TCP-1-delta / CCT-delta / Stimulator of TAR RNA-binding


Mass: 57996.113 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CCT4, CCTD, SRB / Cell line (production host): HEK293F / Production host: Homo sapiens (human) / References: UniProt: P50991
#7: Protein T-complex protein 1 subunit beta / TCP-1-beta / CCT-beta


Mass: 57567.141 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CCT2, 99D8.1, CCTB / Cell line (production host): HEK293F / Production host: Homo sapiens (human) / References: UniProt: P78371
#8: Protein T-complex protein 1 subunit alpha / TCP-1-alpha / CCT-alpha


Mass: 60418.477 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: TCP1, CCT1, CCTA / Cell line (production host): HEK293F / Production host: Homo sapiens (human) / References: UniProt: P17987

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Non-polymers , 4 types, 64 molecules

#9: Chemical
ChemComp-ADP / ADENOSINE-5'-DIPHOSPHATE / Adenosine diphosphate


Mass: 427.201 Da / Num. of mol.: 16 / Source method: obtained synthetically / Formula: C10H15N5O10P2 / Feature type: SUBJECT OF INVESTIGATION / Comment: ADP, energy-carrying molecule*YM
#10: Chemical
ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 16 / Source method: obtained synthetically / Formula: Mg / Feature type: SUBJECT OF INVESTIGATION
#11: Chemical
ChemComp-AF3 / ALUMINUM FLUORIDE / Aluminium fluoride


Mass: 83.977 Da / Num. of mol.: 16 / Source method: obtained synthetically / Formula: AlF3 / Feature type: SUBJECT OF INVESTIGATION
#12: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 16 / Source method: isolated from a natural source / Formula: H2O

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Details

Has ligand of interestY

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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 TRiC-ADP-AlFx / Type: COMPLEX / Entity ID: #1-#8 / Source: NATURAL
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Homo sapiens (human) / Cell: HEK293F
Buffer solutionpH: 7.5
Buffer componentConc.: 50 mM / Name: sodium Chloride / Formula: NaClSodium chloride
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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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 / Nominal defocus max: 2500 nm / Nominal defocus min: 800 nm
Image recordingElectron dose: 38 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 62412 / Symmetry type: POINT

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