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- EMDB-27825: Cryo-EM structure of the endogenous core TIM23 complex from S. ce... -

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Basic information

Entry
Database: EMDB / ID: EMD-27825
TitleCryo-EM structure of the endogenous core TIM23 complex from S. cerevisiae
Map dataUnsharpened map
Sample
  • Complex: Core TIM23 complex (endogenous)
    • Protein or peptide: Mitochondrial import inner membrane translocase subunit TIM17
    • Protein or peptide: Mitochondrial import inner membrane translocase subunit TIM23
    • Protein or peptide: Mitochondrial import inner membrane translocase subunit TIM44
    • Protein or peptide: Antibody Fab fragment light chain
    • Protein or peptide: Antibody Fab fragment heavy chain
  • Ligand: CARDIOLIPIN
  • Ligand: PHOSPHATIDYLETHANOLAMINE
KeywordsTRANSLOCASE-IMMUNE SYSTEM complex
Function / homology
Function and homology information


TIM23 mitochondrial import inner membrane translocase complex / protein import into mitochondrial matrix / protein transmembrane transporter activity / protein-folding chaperone binding / mitochondrial inner membrane
Similarity search - Function
Mitochondrial inner membrane translocase complex, subunit Tim17 / Mitochondrial inner membrane translocase complex, subunit Tim23 / Tim23-like / Mitochondrial import inner membrane translocase subunit Tim44 / Tim44-like / Tim44-like domain / Tim44-like domain / Tim44 / Tim17/Tim22/Tim23/Pmp24 family / NTF2-like domain superfamily
Similarity search - Domain/homology
Mitochondrial import inner membrane translocase subunit TIM17 / Mitochondrial import inner membrane translocase subunit TIM44 / Mitochondrial import inner membrane translocase subunit TIM23
Similarity search - Component
Biological speciesSaccharomyces cerevisiae (brewer's yeast) / Mus musculus (house mouse)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.9 Å
AuthorsSim SI / Park E
Funding support United States, 1 items
OrganizationGrant numberCountry
Other private United States
CitationJournal: Nature / Year: 2023
Title: Structural basis of mitochondrial protein import by the TIM23 complex.
Authors: Sue Im Sim / Yuanyuan Chen / Diane L Lynch / James C Gumbart / Eunyong Park /
Abstract: Mitochondria import nearly all of their approximately 1,000-2,000 constituent proteins from the cytosol across their double-membrane envelope. Genetic and biochemical studies have shown that the ...Mitochondria import nearly all of their approximately 1,000-2,000 constituent proteins from the cytosol across their double-membrane envelope. Genetic and biochemical studies have shown that the conserved protein translocase, termed the TIM23 complex, mediates import of presequence-containing proteins (preproteins) into the mitochondrial matrix and inner membrane. Among about ten different subunits of the TIM23 complex, the essential multipass membrane protein Tim23, together with the evolutionarily related protein Tim17, has long been postulated to form a protein-conducting channel. However, the mechanism by which these subunits form a translocation path in the membrane and enable the import process remains unclear due to a lack of structural information. Here we determined the cryo-electron microscopy structure of the core TIM23 complex (heterotrimeric Tim17-Tim23-Tim44) from Saccharomyces cerevisiae. Contrary to the prevailing model, Tim23 and Tim17 themselves do not form a water-filled channel, but instead have separate, lipid-exposed concave cavities that face in opposite directions. Our structural and biochemical analyses show that the cavity of Tim17, but not Tim23, forms the protein translocation path, whereas Tim23 probably has a structural role. The results further suggest that, during translocation of substrate polypeptides, the nonessential subunit Mgr2 seals the lateral opening of the Tim17 cavity to facilitate the translocation process. We propose a new model for the TIM23-mediated protein import and sorting mechanism, a central pathway in mitochondrial biogenesis.
History
DepositionAug 10, 2022-
Header (metadata) releaseJun 21, 2023-
Map releaseJun 21, 2023-
UpdateOct 4, 2023-
Current statusOct 4, 2023Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_27825.png

Downloads & links

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Map

FileDownload / File: emd_27825.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationUnsharpened map
Voxel sizeX=Y=Z: 1.05 Å
Density
Contour LevelBy AUTHOR: 0.2
Minimum - Maximum-0.32682124 - 1.0412611
Average (Standard dev.)0.0032777665 (±0.02723845)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 268.8 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: Sharpened map

Fileemd_27825_additional_1.map
AnnotationSharpened map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map 1

Fileemd_27825_half_map_1.map
AnnotationHalf map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map 2

Fileemd_27825_half_map_2.map
AnnotationHalf map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Core TIM23 complex (endogenous)

EntireName: Core TIM23 complex (endogenous)
Components
  • Complex: Core TIM23 complex (endogenous)
    • Protein or peptide: Mitochondrial import inner membrane translocase subunit TIM17
    • Protein or peptide: Mitochondrial import inner membrane translocase subunit TIM23
    • Protein or peptide: Mitochondrial import inner membrane translocase subunit TIM44
    • Protein or peptide: Antibody Fab fragment light chain
    • Protein or peptide: Antibody Fab fragment heavy chain
  • Ligand: CARDIOLIPIN
  • Ligand: PHOSPHATIDYLETHANOLAMINE

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Supramolecule #1: Core TIM23 complex (endogenous)

SupramoleculeName: Core TIM23 complex (endogenous) / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#5
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)

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Macromolecule #1: Mitochondrial import inner membrane translocase subunit TIM17

MacromoleculeName: Mitochondrial import inner membrane translocase subunit TIM17
type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Molecular weightTheoretical: 16.602156 KDa
SequenceString:
MSADHSRDPC PIVILNDFGG AFAMGAIGGV VWHGIKGFRN SPLGERGSGA MSAIKARAPV LGGNFGVWGG LFSTFDCAVK AVRKREDPW NAIIAGFFTG GALAVRGGWR HTRNSSITCA CLLGVIEGVG LMFQRYAAWQ AKPMAPPLPE APSSQPLQA

UniProtKB: Mitochondrial import inner membrane translocase subunit TIM17

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Macromolecule #2: Mitochondrial import inner membrane translocase subunit TIM23

MacromoleculeName: Mitochondrial import inner membrane translocase subunit TIM23
type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Molecular weightTheoretical: 23.266527 KDa
SequenceString: MSWLFGDKTP TDDANAAVGG QDTTKPKELS LKQSLGFEPN INNIISGPGG MHVDTARLHP LAGLDKGVEY LDLEEEQLSS LEGSQGLIP SRGWTDDLCY GTGAVYLLGL GIGGFSGMMQ GLQNIPPNSP GKLQLNTVLN HITKRGPFLG NNAGILALSY N IINSTIDA ...String:
MSWLFGDKTP TDDANAAVGG QDTTKPKELS LKQSLGFEPN INNIISGPGG MHVDTARLHP LAGLDKGVEY LDLEEEQLSS LEGSQGLIP SRGWTDDLCY GTGAVYLLGL GIGGFSGMMQ GLQNIPPNSP GKLQLNTVLN HITKRGPFLG NNAGILALSY N IINSTIDA LRGKHDTAGS IGAGALTGAL FKSSKGLKPM GYSSAMVAAA CAVWCSVKKR LLEK

UniProtKB: Mitochondrial import inner membrane translocase subunit TIM23

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Macromolecule #3: Mitochondrial import inner membrane translocase subunit TIM44

MacromoleculeName: Mitochondrial import inner membrane translocase subunit TIM44
type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Molecular weightTheoretical: 48.933418 KDa
SequenceString: MHRSTFIRTS GTSSRTLTAR YRSQYTGLLV ARVLFSTSTT RAQGGNPRSP LQIFRDTFKK EWEKSQELQE NIKTLQDASG KLGESEAYK KAREAYLKAQ RGSTIVGKTL KKTGETMEHI ATKAWESELG KNTRKAAAAT AKKLDESFEP VRQTKIYKEV S EVIDDGES ...String:
MHRSTFIRTS GTSSRTLTAR YRSQYTGLLV ARVLFSTSTT RAQGGNPRSP LQIFRDTFKK EWEKSQELQE NIKTLQDASG KLGESEAYK KAREAYLKAQ RGSTIVGKTL KKTGETMEHI ATKAWESELG KNTRKAAAAT AKKLDESFEP VRQTKIYKEV S EVIDDGES SRYGGFITKE QRRLKRERDL ASGKRHRAVK SNEDAGTAVV ATNIESKESF GKKVEDFKEK TVVGRSIQSL KN KLWDESE NPLIVVMRKI TNKVGGFFAE TESSRVYSQF KLMDPTFSNE SFTRHLREYI VPEILEAYVK GDVKVLKKWF SEA PFNVYA AQQKIFKEQD VYADGRILDI RGVEIVSAKL LAPQDIPVLV VGCRAQEINL YRKKKTGEIA AGDEANILMS SYAM VFTRD PEQIDDDETE GWKILEFVRG GSRQFT

UniProtKB: Mitochondrial import inner membrane translocase subunit TIM44

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Macromolecule #4: Antibody Fab fragment light chain

MacromoleculeName: Antibody Fab fragment light chain / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mus musculus (house mouse)
Molecular weightTheoretical: 26.173201 KDa
Recombinant expressionOrganism: Mus musculus (house mouse)
SequenceString: MEKDTLLLWV LLLWVPGSTG DIVLTQSPAS LAVSLGQRAT ISCRASESVD IYGISFMNWF QQKPGQPPKL LIYATSNQGS GVPARFSGS GSGTDFSLNI HPMEEDDTAM YFCQQSKEVP RTFGGGTKLE IKRADAAPTV SIFPPSSEQL TSGGASVVCF L NNFYPKDI ...String:
MEKDTLLLWV LLLWVPGSTG DIVLTQSPAS LAVSLGQRAT ISCRASESVD IYGISFMNWF QQKPGQPPKL LIYATSNQGS GVPARFSGS GSGTDFSLNI HPMEEDDTAM YFCQQSKEVP RTFGGGTKLE IKRADAAPTV SIFPPSSEQL TSGGASVVCF L NNFYPKDI NVKWKIDGSE RQNGVLNSWT DQDSKDSTYS MSSTLTLTKD EYERHNSYTC EATHKTSTSP IVKSFNRNEC

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Macromolecule #5: Antibody Fab fragment heavy chain

MacromoleculeName: Antibody Fab fragment heavy chain / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mus musculus (house mouse)
Molecular weightTheoretical: 25.49499 KDa
Recombinant expressionOrganism: Mus musculus (house mouse)
SequenceString: MAVLVLLLCL VTFPSCVLSQ VQLKQSGPGL VQPSQSLSIT CTVSGFSLTT YGVHWVRQSP GKGLEWLGVM WRGGSTDFNA AFMSRLSIT KDNSKSQVFF KMNSLQADDT AIYYCARYGN YDAMDYWGQG TSVTVSSAKT TPPSVYPLAP GSAAQTNSMV T LGCLVKGY ...String:
MAVLVLLLCL VTFPSCVLSQ VQLKQSGPGL VQPSQSLSIT CTVSGFSLTT YGVHWVRQSP GKGLEWLGVM WRGGSTDFNA AFMSRLSIT KDNSKSQVFF KMNSLQADDT AIYYCARYGN YDAMDYWGQG TSVTVSSAKT TPPSVYPLAP GSAAQTNSMV T LGCLVKGY FPEPVTVTWN SGSLSSGVHT FPAVLQSDLY TLSSSVTVPS SPRPSETVTC NVAHPASSTK VDKKIVPRDC

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Macromolecule #6: CARDIOLIPIN

MacromoleculeName: CARDIOLIPIN / type: ligand / ID: 6 / Number of copies: 1 / Formula: CDL
Molecular weightTheoretical: 1.464043 KDa
Chemical component information

ChemComp-CDL:
CARDIOLIPIN / phospholipid*YM

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Macromolecule #7: PHOSPHATIDYLETHANOLAMINE

MacromoleculeName: PHOSPHATIDYLETHANOLAMINE / type: ligand / ID: 7 / Number of copies: 1 / Formula: PTY
Molecular weightTheoretical: 734.039 Da
Chemical component information

ChemComp-PTY:
PHOSPHATIDYLETHANOLAMINE / phospholipid*YM

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 7.5
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 400 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 40 sec. / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 0.039 kPa
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 1.6 µm / Nominal defocus min: 0.8 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 1251068
Startup modelType of model: INSILICO MODEL
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 2.15) / Number images used: 214281
Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: NOT APPLICABLE
Final 3D classificationSoftware - Name: cryoSPARC (ver. 2.15)
FSC plot (resolution estimation)

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