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- EMDB-21118: Mouse retromer (VPS26/VPS35/VPS29) chain interface II (VPS26/VPS2... -

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Basic information

Entry
Database: EMDB / ID: EMD-21118
TitleMouse retromer (VPS26/VPS35/VPS29) chain interface II (VPS26/VPS26 dimer)
Map dataMouse retromer (VPS26/VPS35/VPS29) chain interface II (VPS26/VPS26 dimer)
Sample
  • Complex: Mouse retromer (VPS26/VPS35/VPS29) chain interface II (VPS26/VPS26 dimer)
Biological speciesMus musculus (house mouse)
Methodsingle particle reconstruction / cryo EM / Resolution: 16.9 Å
AuthorsKendall AK / Jackson LP
Funding support1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)
CitationJournal: Structure / Year: 2020
Title: Mammalian Retromer Is an Adaptable Scaffold for Cargo Sorting from Endosomes.
Authors: Amy K Kendall / Boyang Xie / Peng Xu / Jue Wang / Rodger Burcham / Meredith N Frazier / Elad Binshtein / Hui Wei / Todd R Graham / Terunaga Nakagawa / Lauren P Jackson /
Abstract: Metazoan retromer (VPS26/VPS35/VPS29) associates with sorting nexins on endosomal tubules to sort proteins to the trans-Golgi network or plasma membrane. Mechanisms of metazoan retromer assembly ...Metazoan retromer (VPS26/VPS35/VPS29) associates with sorting nexins on endosomal tubules to sort proteins to the trans-Golgi network or plasma membrane. Mechanisms of metazoan retromer assembly remain undefined. We combine single-particle cryoelectron microscopy with biophysical methods to uncover multiple oligomer structures. 2D class averages reveal mammalian heterotrimers; dimers of trimers; tetramers of trimers; and flat chains. These species are further supported by biophysical solution studies. We provide reconstructions of all species, including key sub-structures (∼5 Å resolution). Local resolution variation suggests that heterotrimers and dimers adopt multiple conformations. Our structures identify a flexible, highly conserved electrostatic dimeric interface formed by VPS35 subunits. We generate structure-based mutants to disrupt this interface in vitro. Equivalent mutations in yeast demonstrate a mild cargo-sorting defect. Our data suggest the metazoan retromer is an adaptable and plastic scaffold that accommodates interactions with different sorting nexins to sort multiple cargoes from endosomes their final destinations.
History
DepositionDec 13, 2019-
Header (metadata) releaseJan 29, 2020-
Map releaseFeb 19, 2020-
UpdateApr 15, 2020-
Current statusApr 15, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0053
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.0053
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_21118.png

Downloads & links

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Map

FileDownload / File: emd_21118.map.gz / Format: CCP4 / Size: 325 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationMouse retromer (VPS26/VPS35/VPS29) chain interface II (VPS26/VPS26 dimer)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.1 Å/pix.
x 440 pix.
= 482.24 Å		1.1 Å/pix.
x 440 pix.
= 482.24 Å		1.1 Å/pix.
x 440 pix.
= 482.24 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.096 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0053 / Movie #1: 0.0053
Minimum - Maximum-0.011211446 - 0.020573685
Average (Standard dev.)0.0000684930 (±0.00074695674)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions440440440
Spacing440440440
CellA=B=C: 482.24 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.0961.0961.096
M x/y/z440440440
origin x/y/z0.0000.0000.000
length x/y/z482.240482.240482.240
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ400400400
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS440440440
D min/max/mean-0.0110.0210.000

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Supplemental data

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Sample components

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Entire : Mouse retromer (VPS26/VPS35/VPS29) chain interface II (VPS26/VPS2...

EntireName: Mouse retromer (VPS26/VPS35/VPS29) chain interface II (VPS26/VPS26 dimer)
Components
  • Complex: Mouse retromer (VPS26/VPS35/VPS29) chain interface II (VPS26/VPS26 dimer)

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Supramolecule #1: Mouse retromer (VPS26/VPS35/VPS29) chain interface II (VPS26/VPS2...

SupramoleculeName: Mouse retromer (VPS26/VPS35/VPS29) chain interface II (VPS26/VPS26 dimer)
type: complex / ID: 1 / Parent: 0
Source (natural)Organism: Mus musculus (house mouse)
Recombinant expressionOrganism: Escherichia coli (E. coli)

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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

Concentration0.1 mg/mL
BufferpH: 8.2
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 71.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionResolution.type: BY AUTHOR / Resolution: 16.9 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 13782
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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