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- PDB-8xks: The cryo-EM structure of Orf2971-FtsHi motor complex -

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Basic information

Entry
Database: PDB / ID: 8xks
TitleThe cryo-EM structure of Orf2971-FtsHi motor complex
Components
  • (AAA+ ATPase domain-containing ...) x 2
  • 4Fe-4S ferredoxin-type domain-containing protein
  • ADP-ribosylglycohydrolase
  • Acyl carrier protein
  • Ctap1
  • Fatty acid desaturase domain-containing protein
  • FaxL
  • Fhl2
  • Flagellar associated protein
  • Moc13
  • Moc25
  • Moc29
  • Moc31
  • Moc34
  • Moc45
  • PcyA1
  • Tic22
  • Uncharacterized 341.7 kDa protein in psbD-psbC intergenic region
KeywordsPROTEIN TRANSPORT / Chlamydomonas reinhardtii / protein transportation / chloroplast / lipid synthesis / Enzyme
Function / homology
Function and homology information


fatty acid transmembrane transporter activity / chloroplast inner membrane / phytochromobilin biosynthetic process / oxidoreductase activity, acting on the CH-CH group of donors, iron-sulfur protein as acceptor / poly(ADP-ribose) glycohydrolase / cobalt ion binding / ATP-dependent peptidase activity / chloroplast thylakoid membrane / acyl carrier activity / fatty acid transport ...fatty acid transmembrane transporter activity / chloroplast inner membrane / phytochromobilin biosynthetic process / oxidoreductase activity, acting on the CH-CH group of donors, iron-sulfur protein as acceptor / poly(ADP-ribose) glycohydrolase / cobalt ion binding / ATP-dependent peptidase activity / chloroplast thylakoid membrane / acyl carrier activity / fatty acid transport / chloroplast / metalloendopeptidase activity / protein transport / oxidoreductase activity / hydrolase activity / ATP hydrolysis activity / proteolysis / ATP binding / membrane
Similarity search - Function
Tic22-like / Tic22-like family / Fatty acid desaturase / Acyl carrier protein, chloroplastic / Ferredoxin-dependent bilin reductase / Ferredoxin-dependent bilin reductase / ADP-ribosylation/Crystallin J1 / ADP-ribosylglycohydrolase / ADP-ribosylation/Crystallin J1 superfamily / : ...Tic22-like / Tic22-like family / Fatty acid desaturase / Acyl carrier protein, chloroplastic / Ferredoxin-dependent bilin reductase / Ferredoxin-dependent bilin reductase / ADP-ribosylation/Crystallin J1 / ADP-ribosylglycohydrolase / ADP-ribosylation/Crystallin J1 superfamily / : / Peptidase M41 / Peptidase M41-like / Peptidase family M41 / : / Acyl carrier protein (ACP) / AAA ATPase, AAA+ lid domain / AAA+ lid domain / ATPase, AAA-type, conserved site / AAA-protein family signature. / Phosphopantetheine attachment site / ACP-like superfamily / Carrier protein (CP) domain profile. / Phosphopantetheine binding ACP domain / ATPase family associated with various cellular activities (AAA) / ATPase, AAA-type, core / ATPases associated with a variety of cellular activities / AAA+ ATPase domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
ARACHIDONIC ACID / DIACYL GLYCEROL / DIGALACTOSYL DIACYL GLYCEROL (DGDG) / 1,2-DISTEAROYL-MONOGALACTOSYL-DIGLYCERIDE / Chem-SQD / Uncharacterized protein / Uncharacterized protein / Uncharacterized protein / 4Fe-4S ferredoxin-type domain-containing protein / Flagellar associated protein ...ARACHIDONIC ACID / DIACYL GLYCEROL / DIGALACTOSYL DIACYL GLYCEROL (DGDG) / 1,2-DISTEAROYL-MONOGALACTOSYL-DIGLYCERIDE / Chem-SQD / Uncharacterized protein / Uncharacterized protein / Uncharacterized protein / 4Fe-4S ferredoxin-type domain-containing protein / Flagellar associated protein / AAA+ ATPase domain-containing protein / ADP-ribosylhydrolase ARH3 / Uncharacterized protein / Fatty acid desaturase domain-containing protein / Uncharacterized protein / AAA+ ATPase domain-containing protein / Uncharacterized protein / Uncharacterized protein / Uncharacterized protein / Uncharacterized 341.7 kDa protein in psbD-psbC intergenic region / Acyl carrier protein
Similarity search - Component
Biological speciesChlamydomonas reinhardtii (plant)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.2 Å
AuthorsWang, N. / Li, M.
Funding support China, 1items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC) China
CitationJournal: Mol Plant / Year: 2024
Title: Architecture of the ATP-driven motor for protein import into chloroplasts.
Authors: Ning Wang / Jiale Xing / Xiaodong Su / Junting Pan / Hui Chen / Lifang Shi / Long Si / Wenqiang Yang / Mei Li /
Abstract: Thousands of nuclear-encoded proteins are transported into chloroplasts through the TOC-TIC translocon that spans the chloroplast envelope membranes. A motor complex pulls the translocated proteins ...Thousands of nuclear-encoded proteins are transported into chloroplasts through the TOC-TIC translocon that spans the chloroplast envelope membranes. A motor complex pulls the translocated proteins out of the TOC-TIC complex into the chloroplast stroma by hydrolyzing ATP. The Orf2971-FtsHi complex has been suggested to serve as the ATP-hydrolyzing motor in Chlamydomonas reinhardtii, but little is known about its architecture and assembly. Here, we report the 3.2-Å resolution structure of the Chlamydomonas Orf2971-FtsHi complex. The 20-subunit complex spans the chloroplast inner envelope, with two bulky modules protruding into the intermembrane space and stromal matrix. Six subunits form a hetero-hexamer that potentially provides the pulling force through ATP hydrolysis. The remaining subunits, including potential enzymes/chaperones, likely facilitate the complex assembly and regulate its proper function. Taken together, our results provide the structural foundation for a mechanistic understanding of chloroplast protein translocation.
History
DepositionDec 24, 2023Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Dec 4, 2024Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Ctap1
B: Fhl2
C: 4Fe-4S ferredoxin-type domain-containing protein
D: AAA+ ATPase domain-containing protein
E: Uncharacterized 341.7 kDa protein in psbD-psbC intergenic region
F: AAA+ ATPase domain-containing protein
G: AAA+ ATPase domain-containing protein
H: Flagellar associated protein
I: Tic22
J: FaxL
K: Fatty acid desaturase domain-containing protein
L: Moc25
M: Moc29
N: Moc34
O: Moc45
P: PcyA1
Q: ADP-ribosylglycohydrolase
R: Acyl carrier protein
S: Moc13
T: Moc31
hetero molecules


Theoretical massNumber of molelcules
Total (without water)1,509,08129
Polymers1,503,75320
Non-polymers5,3289
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 , 17 types, 17 molecules ABCEHIJKLMNOPQRST

#1: Protein Ctap1


Mass: 107778.039 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant)
#2: Protein Fhl2


Mass: 109355.469 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant)
#3: Protein 4Fe-4S ferredoxin-type domain-containing protein / Moc50


Mass: 50682.551 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: A0A2K3DGW6
#5: Protein Uncharacterized 341.7 kDa protein in psbD-psbC intergenic region / ORF2971 / ORFB


Mass: 342116.750 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: Q32065
#7: Protein Flagellar associated protein / Ctap7


Mass: 71675.305 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: A0A2K3DHV6
#8: Protein Tic22


Mass: 36869.215 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: A0A2K3CYB6
#9: Protein FaxL


Mass: 39091.016 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: A0A2K3DM81
#10: Protein Fatty acid desaturase domain-containing protein / FAD6a


Mass: 70334.234 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: A0A2K3DQ46
#11: Protein Moc25


Mass: 29084.951 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: A0A2K3CY83
#12: Protein Moc29


Mass: 34185.254 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: A0A2K3CZ99
#13: Protein Moc34


Mass: 34163.977 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: A0A2K3E633
#14: Protein Moc45


Mass: 48618.184 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: A0A2K3DVJ8
#15: Protein PcyA1


Mass: 61329.266 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: A8HUP3
#16: Protein ADP-ribosylglycohydrolase / ArhL


Mass: 51410.547 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: A0A2K3DLI2
#17: Protein Acyl carrier protein / ACP2


Mass: 12519.384 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: Q6UKY5
#18: Protein Moc13


Mass: 15265.472 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: A8ISA4
#19: Protein Moc31


Mass: 31085.385 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant)

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AAA+ ATPase domain-containing ... , 2 types, 3 molecules DFG

#4: Protein AAA+ ATPase domain-containing protein / Fhl1


Mass: 128535.375 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: A0A2K3DZD9
#6: Protein AAA+ ATPase domain-containing protein / Fhl3a


Mass: 114826.094 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Chlamydomonas reinhardtii (plant) / References: UniProt: A0A2K3DLF7

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Sugars , 1 types, 2 molecules

#22: Sugar ChemComp-DGD / DIGALACTOSYL DIACYL GLYCEROL (DGDG)


Type: saccharide / Mass: 949.299 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C51H96O15

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Non-polymers , 5 types, 7 molecules

#20: Chemical ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Zn / Feature type: SUBJECT OF INVESTIGATION
#21: Chemical ChemComp-DGA / DIACYL GLYCEROL


Mass: 625.018 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C39H76O5 / Feature type: SUBJECT OF INVESTIGATION
#23: Chemical ChemComp-LMG / 1,2-DISTEAROYL-MONOGALACTOSYL-DIGLYCERIDE


Mass: 787.158 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C45H86O10
#24: Chemical ChemComp-ACD / ARACHIDONIC ACID


Mass: 304.467 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C20H32O2 / Feature type: SUBJECT OF INVESTIGATION
#25: Chemical ChemComp-SQD / 1,2-DI-O-ACYL-3-O-[6-DEOXY-6-SULFO-ALPHA-D-GLUCOPYRANOSYL]-SN-GLYCEROL / SULFOQUINOVOSYLDIACYLGLYCEROL


Mass: 795.116 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C41H78O12S

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Details

Has ligand of interestY
Has protein modificationY

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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: The complex of Orf2971-FtsHi motor / Type: COMPLEX / Entity ID: #1-#19 / Source: NATURAL
Molecular weightExperimental value: NO
Source (natural)Organism: Chlamydomonas reinhardtii (plant)
Buffer solutionpH: 7.4
SpecimenConc.: 1.2 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 400 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationCryogen name: ETHANE

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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: DIFFRACTION / Nominal defocus max: 2000 nm / Nominal defocus min: 1200 nm
Specimen holderCryogen: NITROGEN
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

EM softwareName: PHENIX / Version: 1.20.1_4487: / Category: model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 127613 / Symmetry type: POINT
Atomic model buildingProtocol: BACKBONE TRACE / Space: REAL
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0169087
ELECTRON MICROSCOPYf_angle_d1.03893764
ELECTRON MICROSCOPYf_dihedral_angle_d6.1269567
ELECTRON MICROSCOPYf_chiral_restr0.05210304
ELECTRON MICROSCOPYf_plane_restr0.00812118

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