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- EMDB-63981: Cryo-EM structure of complex III2 of mammalian respiratory superc... -

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Basic information

Entry
Database: EMDB / ID: EMD-63981
TitleCryo-EM structure of complex III2 of mammalian respiratory supercomplex in vesicles
Map data
Sample
  • Complex: The complex III2 of the respiratory supercomplex from pig's heart mitochondria
KeywordsMembrane protein / Vesicle / Methodology / mammalian cells / mitochondrion / respiratory supercomplex / native structure
Biological speciesSus scrofa (pig)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.0 Å
AuthorsLiu H / Dang S
Funding support Hong Kong, 1 items
OrganizationGrant numberCountry
The University Grants Committee, Research Grants Council (RGC) Hong Kong
CitationJournal: Proc Natl Acad Sci U S A / Year: 2025
Title: Capturing the native structure of membrane proteins using vesicles.
Authors: Hang Liu / Chun Mong Tse / Shangyu Dang /
Abstract: Membrane proteins play crucial roles in numerous biological processes and are important drug targets. However, structural studies of membrane proteins often rely on solubilization with detergents, ...Membrane proteins play crucial roles in numerous biological processes and are important drug targets. However, structural studies of membrane proteins often rely on solubilization with detergents, which may not accurately reflect their native states in a cellular context. Additionally, identifying suitable detergents for individual membrane proteins can be a detailed and time-consuming process. Here, we developed a vesicle-based method that preserves the native lipid environment for subsequent structural and functional studies. Using the bacterial multidrug efflux transporter AcrB as an example, we isolated AcrB-containing vesicles and determined its cryo-EM structure with all protomers in a loose (L) state at 3.88 Å by incorporating our micrograph-based sorting strategy. Notably, compared to the L-state AcrB in liposomes and nanoparticles, the exterior transmembrane helices (TMs) in our map exhibited superior quality, featuring a continuous and clear representation of lα, which is positioned horizontally within the lipid bilayer. We further expanded our method by identifying endogenous membrane proteins, including F-ATPase and respiratory complexes, in vesicles generated using mitochondria from pig hearts. The high-resolution structure of respiratory complex III in vesicles revealed a shared subunit nine between two monomers. Briefly, our method presents a promising and straightforward approach for studying the structure and function of membrane proteins in their native environment, eliminating the need for detergent screening and protein purification.
History
DepositionMar 31, 2025-
Header (metadata) releaseAug 27, 2025-
Map releaseAug 27, 2025-
UpdateSep 24, 2025-
Current statusSep 24, 2025Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_63981.png

Downloads & links

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Map

FileDownload / File: emd_63981.map.gz / Format: CCP4 / Size: 91.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

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AxesZ (Sec.)Y (Row.)X (Col.)
1.06 Å/pix.
x 288 pix.
= 305.28 Å		1.06 Å/pix.
x 288 pix.
= 305.28 Å		1.06 Å/pix.
x 288 pix.
= 305.28 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1.06 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.1
Minimum - Maximum-0.04102156 - 2.430019
Average (Standard dev.)0.0028411665 (±0.039212026)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions288288288
Spacing288288288
CellA=B=C: 305.27997 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_63981_msk_1.map
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Half map: #1

Fileemd_63981_half_map_1.map
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Half map: #2

Fileemd_63981_half_map_2.map
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Sample components

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Entire : The complex III2 of the respiratory supercomplex from pig's heart...

EntireName: The complex III2 of the respiratory supercomplex from pig's heart mitochondria
Components
  • Complex: The complex III2 of the respiratory supercomplex from pig's heart mitochondria

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Supramolecule #1: The complex III2 of the respiratory supercomplex from pig's heart...

SupramoleculeName: The complex III2 of the respiratory supercomplex from pig's heart mitochondria
type: complex / ID: 1 / Parent: 0
Source (natural)Organism: Sus scrofa (pig) / Organ: heart / Tissue: muscle / Organelle: mitochondria

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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
VitrificationCryogen name: ETHANE
DetailsVesicles of mitochondria membrane

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average exposure time: 3.74 sec. / 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 / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: INSILICO MODEL
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.0 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 42832
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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