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- EMDB-20590: CryoEM reconstruction of membrane-bound ESCRT-III filament compos... -

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Basic information

Entry
Database: EMDB / ID: EMD-20590
TitleCryoEM reconstruction of membrane-bound ESCRT-III filament composed of CHMP1B only
Map dataRELION Refine3D filtered map with helical symmetry imposed on the central 40%
Sample
  • Complex: membrane-bound ESCRT-III copolymer filament composed of CHMP1B only
    • Protein or peptide: Charged multivesicular body protein 1b
Keywordsmembrane remodeling / membrane-bound protein filament / ESCRT-III / LIPID BINDING PROTEIN
Function / homology
Function and homology information


MIT domain binding / amphisome membrane / multivesicular body-lysosome fusion / vesicle fusion with vacuole / ESCRT III complex disassembly / late endosome to lysosome transport / ESCRT III complex / kinetochore microtubule / endosome transport via multivesicular body sorting pathway / regulation of centrosome duplication ...MIT domain binding / amphisome membrane / multivesicular body-lysosome fusion / vesicle fusion with vacuole / ESCRT III complex disassembly / late endosome to lysosome transport / ESCRT III complex / kinetochore microtubule / endosome transport via multivesicular body sorting pathway / regulation of centrosome duplication / nuclear membrane reassembly / midbody abscission / multivesicular body sorting pathway / membrane coat / membrane fission / plasma membrane repair / late endosome to vacuole transport / multivesicular body membrane / ubiquitin-dependent protein catabolic process via the multivesicular body sorting pathway / multivesicular body assembly / regulation of mitotic spindle assembly / nucleus organization / mitotic metaphase chromosome alignment / viral budding via host ESCRT complex / autophagosome maturation / autophagosome membrane / nuclear pore / multivesicular body / viral budding from plasma membrane / establishment of protein localization / kinetochore / autophagy / protein transport / midbody / endosome membrane / cell division / lysosomal membrane / protein domain specific binding / extracellular exosome / nucleoplasm / identical protein binding / plasma membrane / cytosol
Similarity search - Function
Charged multivesicular body protein 1b
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodhelical reconstruction / cryo EM / Resolution: 6.2 Å
AuthorsNguyen HC / Frost A
Funding support United States, 5 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)P50 AI150464 United States
National Institutes of Health/Office of the DirectorS10OD020054 United States
National Institutes of Health/Office of the DirectorS10OD021741 United States
National Institutes of Health/Office of the Director1S10OD021596-01 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)1DP2GM110772-01 United States
CitationJournal: Nat Struct Mol Biol / Year: 2020
Title: Membrane constriction and thinning by sequential ESCRT-III polymerization.
Authors: Henry C Nguyen / Nathaniel Talledge / John McCullough / Abhimanyu Sharma / Frank R Moss / Janet H Iwasa / Michael D Vershinin / Wesley I Sundquist / Adam Frost /
Abstract: The endosomal sorting complexes required for transport (ESCRTs) mediate diverse membrane remodeling events. These typically require ESCRT-III proteins to stabilize negatively curved membranes; ...The endosomal sorting complexes required for transport (ESCRTs) mediate diverse membrane remodeling events. These typically require ESCRT-III proteins to stabilize negatively curved membranes; however, recent work has indicated that certain ESCRT-IIIs also participate in positive-curvature membrane-shaping reactions. ESCRT-IIIs polymerize into membrane-binding filaments, but the structural basis for negative versus positive membrane remodeling by these proteins remains poorly understood. To learn how certain ESCRT-IIIs shape positively curved membranes, we determined structures of human membrane-bound CHMP1B-only, membrane-bound CHMP1B + IST1, and IST1-only filaments by cryo-EM. Our structures show how CHMP1B first polymerizes into a single-stranded helical filament, shaping membranes into moderate-curvature tubules. Subsequently, IST1 assembles a second strand on CHMP1B, further constricting the membrane tube and reducing its diameter nearly to the fission point. Each step of constriction thins the underlying bilayer, lowering the barrier to membrane fission. Our structures reveal how a two-component, sequential polymerization mechanism drives membrane tubulation, constriction and bilayer thinning.
History
DepositionAug 11, 2019-
Header (metadata) releaseSep 18, 2019-
Map releaseApr 8, 2020-
UpdateMar 20, 2024-
Current statusMar 20, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.01
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.01
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6tz9
  • Surface level: 0.01
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-6tz9
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_20590.png

Downloads & links

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Map

FileDownload / File: emd_20590.map.gz / Format: CCP4 / Size: 129.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationRELION Refine3D filtered map with helical symmetry imposed on the central 40%
Voxel sizeX=Y=Z: 1.345 Å
Density
Contour LevelBy AUTHOR: 0.01 / Movie #1: 0.01
Minimum - Maximum-0.0067385007 - 0.027849544
Average (Standard dev.)0.00023361685 (±0.0036929275)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions324324324
Spacing324324324
CellA=B=C: 435.78 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.3451.3451.345
M x/y/z324324324
origin x/y/z0.0000.0000.000
length x/y/z435.780435.780435.780
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS324324324
D min/max/mean-0.0070.0280.000

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Supplemental data

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Mask #1

Fileemd_20590_msk_1.map
Projections & Slices
AxesZYX

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Additional map: RELION postprocessed map with helical symmetry applied to...

Fileemd_20590_additional_1.map
AnnotationRELION postprocessed map with helical symmetry applied to the entire map
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AxesZYX

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Additional map: RELION Refine3D filtered map with no symmetry applied (C1 map)

Fileemd_20590_additional_2.map
AnnotationRELION Refine3D filtered map with no symmetry applied (C1 map)
Projections & Slices
AxesZYX

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Additional map: RELION Refine3D unfiltered half map1 with no symmetry...

Fileemd_20590_additional_3.map
AnnotationRELION Refine3D unfiltered half map1 with no symmetry applied (C1 half1 map)
Projections & Slices
AxesZYX

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Additional map: RELION Refine3D unfiltered half map2 with no symmetry...

Fileemd_20590_additional_4.map
AnnotationRELION Refine3D unfiltered half map2 with no symmetry applied (C1 half2 map)
Projections & Slices
AxesZYX

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Histogram

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Half map: RELION Refine3D unfiltered half map1 with helical symmetry...

Fileemd_20590_half_map_1.map
AnnotationRELION Refine3D unfiltered half map1 with helical symmetry imposed on the central 40%
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: RELION Refine3D unfiltered half map2 with helical symmetry...

Fileemd_20590_half_map_2.map
AnnotationRELION Refine3D unfiltered half map2 with helical symmetry imposed on the central 40%
Projections & Slices
AxesZYX

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Slices (1/2)
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Histogram (log scale)

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Sample components

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Entire : membrane-bound ESCRT-III copolymer filament composed of CHMP1B only

EntireName: membrane-bound ESCRT-III copolymer filament composed of CHMP1B only
Components
  • Complex: membrane-bound ESCRT-III copolymer filament composed of CHMP1B only
    • Protein or peptide: Charged multivesicular body protein 1b

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Supramolecule #1: membrane-bound ESCRT-III copolymer filament composed of CHMP1B only

SupramoleculeName: membrane-bound ESCRT-III copolymer filament composed of CHMP1B only
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Charged multivesicular body protein 1b

MacromoleculeName: Charged multivesicular body protein 1b / type: protein_or_peptide / ID: 1 / Number of copies: 26 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 22.140354 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MSNMEKHLFN LKFAAKELSR SAKKCDKEEK AEKAKIEKAI QKGNMEVARI HAENAIRQKN QAVNFLRMSA RVDAVAARVQ TAVTMGKVT KSMAGVVKSM DATLKTMNLE KISALMDKFE HQFETLDVQT QQMEDTMSST TTLTTPQNQV DMLLQEMADE A GLDLNMEL ...String:
MSNMEKHLFN LKFAAKELSR SAKKCDKEEK AEKAKIEKAI QKGNMEVARI HAENAIRQKN QAVNFLRMSA RVDAVAARVQ TAVTMGKVT KSMAGVVKSM DATLKTMNLE KISALMDKFE HQFETLDVQT QQMEDTMSST TTLTTPQNQV DMLLQEMADE A GLDLNMEL PQGQTGSVGT SVASAEQDEL SQRLARLRDQ V

UniProtKB: Charged multivesicular body protein 1b

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Experimental details

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Structure determination

Methodcryo EM
Processinghelical reconstruction
Aggregation statehelical array

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Sample preparation

BufferpH: 7.4
GridPretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec. / Details: unspecified
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 292 K / Instrument: FEI VITROBOT MARK III
Details: Grids were blotted with Whatman No. 1 filter paper for 4-8 seconds with a 0 mm offset at 19C and 100 percent humidity before plunging into liquid ethane.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Average electron dose: 45.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL / In silico model: Smooth cylinder
Final angle assignmentType: NOT APPLICABLE / Software - Name: RELION
Final reconstructionApplied symmetry - Helical parameters - Δz: 1.86 Å
Applied symmetry - Helical parameters - Δ&Phi: 13.86 °
Applied symmetry - Helical parameters - Axial symmetry: C1 (asymmetric)
Resolution.type: BY AUTHOR / Resolution: 6.2 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION / Number images used: 9661
FSC plot (resolution estimation)

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