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- PDB-7xzi: Cryo-EM structure of TOC-TIC supercomplex from Chlamydomonas rein... -

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Basic information

Entry
Database: PDB / ID: 7xzi
TitleCryo-EM structure of TOC-TIC supercomplex from Chlamydomonas reinhardtii
Components
  • Ctap3
  • Ctap4
  • Ctap5
  • Protein TIC 20
  • Simp2
  • Simp3
  • Tic100
  • Tic15
  • Tic214
  • Tic56
  • Toc34
  • Toc75
  • Toc90
  • Unknown peptide
KeywordsTRANSLOCASE / Complex / membrane protein
Function / homology
Function and homology information


protein import into chloroplast stroma / chloroplast inner membrane / chloroplast outer membrane / chloroplast membrane / Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement / protein transport / membrane => GO:0016020 / hydrolase activity / GTP binding / membrane / metal ion binding
Similarity search - Function
CCB3/YggT / Chloroplast protein import component Tic20 / Protein TIC56 / YGGT family / Chloroplast import apparatus Tic20-like / GYF domain 2 / GYF domain 2 / GTPase GIMA/IAN/Toc / AIG1-type guanine nucleotide-binding (G) domain / AIG1 family ...CCB3/YggT / Chloroplast protein import component Tic20 / Protein TIC56 / YGGT family / Chloroplast import apparatus Tic20-like / GYF domain 2 / GYF domain 2 / GTPase GIMA/IAN/Toc / AIG1-type guanine nucleotide-binding (G) domain / AIG1 family / AIG1-type G domain profile. / Possible plasma membrane-binding motif in junctophilins, PIP-5-kinases and protein kinases. / MORN motif / MORN repeat / Surface antigen D15-like / Bacterial surface antigen (D15) / Omp85 superfamily domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
DODECANE / INOSITOL HEXAKISPHOSPHATE / 1,2-DIPALMITOYL-PHOSPHATIDYL-GLYCEROLE / 1,2-DISTEAROYL-MONOGALACTOSYL-DIGLYCERIDE / HEXADECANE / AIG1-type G domain-containing protein / Flagellar associated protein / Uncharacterized protein / YggT family protein / Translocase of chloroplast 34 homolog, chloroplastic ...DODECANE / INOSITOL HEXAKISPHOSPHATE / 1,2-DIPALMITOYL-PHOSPHATIDYL-GLYCEROLE / 1,2-DISTEAROYL-MONOGALACTOSYL-DIGLYCERIDE / HEXADECANE / AIG1-type G domain-containing protein / Flagellar associated protein / Uncharacterized protein / YggT family protein / Translocase of chloroplast 34 homolog, chloroplastic / Bacterial surface antigen (D15) domain-containing protein / Protein TIC 20 / Uncharacterized protein / Uncharacterized protein / Uncharacterized protein / Uncharacterized protein / Uncharacterized membrane protein ycf78
Similarity search - Component
Biological speciesChlamydomonas reinhardtii (plant)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.77 Å
AuthorsLiu, H. / Li, A.J. / Liu, Z.F.
Funding support China, 4items
OrganizationGrant numberCountry
Chinese Academy of SciencesYSBR-015 China
National Natural Science Foundation of China (NSFC)31925024 China
Chinese Academy of SciencesXDB37020101 China
Ministry of Science and Technology (MoST, China)2017YFA0503702 China
CitationJournal: Nature / Year: 2023
Title: Architecture of chloroplast TOC-TIC translocon supercomplex.
Authors: Hao Liu / Anjie Li / Jean-David Rochaix / Zhenfeng Liu /
Abstract: Chloroplasts rely on the translocon complexes in the outer and inner envelope membranes (the TOC and TIC complexes, respectively) to import thousands of different nuclear-encoded proteins from the ...Chloroplasts rely on the translocon complexes in the outer and inner envelope membranes (the TOC and TIC complexes, respectively) to import thousands of different nuclear-encoded proteins from the cytosol. Although previous studies indicated that the TOC and TIC complexes may assemble into larger supercomplexes, the overall architectures of the TOC-TIC supercomplexes and the mechanism of preprotein translocation are unclear. Here we report the cryo-electron microscopy structure of the TOC-TIC supercomplex from Chlamydomonas reinhardtii. The major subunits of the TOC complex (Toc75, Toc90 and Toc34) and TIC complex (Tic214, Tic20, Tic100 and Tic56), three chloroplast translocon-associated proteins (Ctap3, Ctap4 and Ctap5) and three newly identified small inner-membrane proteins (Simp1-3) have been located in the supercomplex. As the largest protein, Tic214 traverses the inner membrane, the intermembrane space and the outer membrane, connecting the TOC complex with the TIC proteins. An inositol hexaphosphate molecule is located at the Tic214-Toc90 interface and stabilizes their assembly. Four lipid molecules are located within or above an inner-membrane funnel formed by Tic214, Tic20, Simp1 and Ctap5. Multiple potential pathways found in the TOC-TIC supercomplex may support translocation of different substrate preproteins into chloroplasts.
History
DepositionJun 2, 2022Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Jan 11, 2023Provider: repository / Type: Initial release
Revision 1.1Jan 18, 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.pdbx_database_id_DOI / _citation.title / _citation.year / _citation_author.identifier_ORCID / _citation_author.name
Revision 1.2Feb 15, 2023Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _citation_author.identifier_ORCID
Revision 1.3Mar 22, 2023Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation_author.identifier_ORCID

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
3: Ctap3
4: Ctap4
5: Ctap5
7: Toc75
9: Toc90
A: Tic214
B: Protein TIC 20
C: Tic15
D: Simp3
E: Tic100
F: Tic56
G: Toc34
U: Simp2
X: Unknown peptide
hetero molecules


Theoretical massNumber of molelcules
Total (without water)830,92538
Polymers815,14914
Non-polymers15,77624
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: native gel electrophoresis, Blue native PAGE shows the molecular weight of the complex is about 1 MDa., gel filtration, SEC shows the molecular weight of the complex is about 1 MDa.
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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Protein , 13 types, 13 molecules 34579ABCDEFGU

#1: Protein Ctap3


Mass: 51982.613 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: A0A2K3D4W3
#2: Protein Ctap4 / Omp85 domain-containing protein


Mass: 39025.082 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: A8J6H7
#3: Protein Ctap5


Mass: 42710.965 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant)
#4: Protein Toc75 / 75 kDa chloroplast membrane translocon


Mass: 87333.289 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: A8IE32
#5: Protein Toc90 / AIG1-type G domain-containing protein


Mass: 102415.258 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: A0A2K3CR90
#6: Protein Tic214 / Uncharacterized membrane protein ycf78


Mass: 232535.719 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: P36495
#7: Protein Protein TIC 20


Mass: 29545.480 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: A8IZ79
#8: Protein Tic15


Mass: 14662.776 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: A8J1J3
#9: Protein Simp3


Mass: 21185.650 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: A0A2K3E0K3
#10: Protein Tic100


Mass: 105817.695 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: A0A2K3DQY7
#11: Protein Tic56


Mass: 28046.527 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: A8J6R5
#12: Protein Toc34 / Translocase of chloroplast 34 homolog / chloroplastic


Mass: 44598.613 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant)
References: UniProt: A8HYJ1, Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement
#13: Protein Simp2 / Tic13


Mass: 13426.158 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: A8J5D4

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Protein/peptide , 1 types, 1 molecules X

#14: Protein/peptide Unknown peptide


Mass: 1863.077 Da / Num. of mol.: 1 / Source method: isolated from a natural source
Details: Chain X belongs to an unidentified protein, which contains an alpha helix. The input sequence is a tentative one based on the model. The identity of this chain remains to be revealed in the future.
Source: (natural) Chlamydomonas reinhardtii (plant)

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Non-polymers , 6 types, 24 molecules

#15: Chemical
ChemComp-AJP / Digitonin / Digitonin


Mass: 1229.312 Da / Num. of mol.: 5 / Source method: obtained synthetically / Formula: C56H92O29 / Comment: detergent*YM
#16: Chemical
ChemComp-LMG / 1,2-DISTEAROYL-MONOGALACTOSYL-DIGLYCERIDE


Mass: 787.158 Da / Num. of mol.: 5 / Source method: obtained synthetically / Formula: C45H86O10 / Feature type: SUBJECT OF INVESTIGATION
#17: Chemical ChemComp-R16 / HEXADECANE / Hexadecane


Mass: 226.441 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C16H34
#18: Chemical
ChemComp-D12 / DODECANE / Dodecane


Mass: 170.335 Da / Num. of mol.: 7 / Source method: obtained synthetically / Formula: C12H26
#19: Chemical ChemComp-IHP / INOSITOL HEXAKISPHOSPHATE / MYO-INOSITOL HEXAKISPHOSPHATE / INOSITOL 1,2,3,4,5,6-HEXAKISPHOSPHATE / Phytic acid


Mass: 660.035 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C6H18O24P6 / Feature type: SUBJECT OF INVESTIGATION
#20: Chemical
ChemComp-LHG / 1,2-DIPALMITOYL-PHOSPHATIDYL-GLYCEROLE / Phosphatidylglycerol


Mass: 722.970 Da / Num. of mol.: 5 / Source method: obtained synthetically / Formula: C38H75O10P / Feature type: SUBJECT OF INVESTIGATION / Comment: phospholipid*YM

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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: a membrane protein complex / Type: COMPLEX / Entity ID: #1-#14 / Source: NATURAL
Molecular weightValue: 0.812 MDa / Experimental value: YES
Source (natural)Organism: Chlamydomonas reinhardtii (plant)
Buffer solutionpH: 7.5
Buffer component
IDConc.NameFormulaBuffer-ID
150 mMTrisC4H11NO31
2150 mMSodium chlorideNaClSodium chloride1
30.06 %DigitoninC56H92O291
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid type: Quantifoil
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 %

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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: OTHER
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 1800 nm / Nominal defocus min: 1500 nm
Image recordingElectron dose: 50 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K2 QUANTUM (4k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.20.1_4487: / Classification: refinement
EM software
IDNameVersionCategory
4cryoSPARCv3.2CTF correction
7Coot0.9.4model fitting
9PHENIX1.20.1-4487model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 2.77 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 796731 / Num. of class averages: 1 / Symmetry type: POINT
Atomic model buildingProtocol: AB INITIO MODEL / Space: REAL
RefinementHighest resolution: 2.77 Å / Cross valid method: NONE
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00238276
ELECTRON MICROSCOPYf_angle_d0.45351901
ELECTRON MICROSCOPYf_dihedral_angle_d10.55913835
ELECTRON MICROSCOPYf_chiral_restr0.0395619
ELECTRON MICROSCOPYf_plane_restr0.0046606

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