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- EMDB-7141: Glutamate dehydrogenase on a holey carbon grid -

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Basic information

Entry
Database: EMDB / ID: EMD-7141
TitleGlutamate dehydrogenase on a holey carbon grid
Map dataTomogram of GDH single particle
Sample
  • Complex: Glutamate dehydrogenase
Biological speciesunidentified (others)
Methodelectron tomography / cryo EM
AuthorsNoble AN / Dandey VP / Wei H / Brasch J / Chase J / Acharya P / Tan Y / Zhang Z / Kim LY / Scapin G ...Noble AN / Dandey VP / Wei H / Brasch J / Chase J / Acharya P / Tan Y / Zhang Z / Kim LY / Scapin G / Rapp M / Eng ET / Rice WJ / Cheng A / Negro CJ / Shapiro L / Kwong PD / Jeruzalmi D / des Georges A / Potter CS / Carragher B
Funding support United States, 5 items
OrganizationGrant numberCountry
Simons Foundation349247 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM103310 United States
NYSTAR United States
National Institutes of Health/Office of the DirectorS10 OD019994-01 United States
Agouron InstituteF00316 United States
CitationJournal: Elife / Year: 2018
Title: Routine single particle CryoEM sample and grid characterization by tomography.
Authors: Alex J Noble / Venkata P Dandey / Hui Wei / Julia Brasch / Jillian Chase / Priyamvada Acharya / Yong Zi Tan / Zhening Zhang / Laura Y Kim / Giovanna Scapin / Micah Rapp / Edward T Eng / ...Authors: Alex J Noble / Venkata P Dandey / Hui Wei / Julia Brasch / Jillian Chase / Priyamvada Acharya / Yong Zi Tan / Zhening Zhang / Laura Y Kim / Giovanna Scapin / Micah Rapp / Edward T Eng / William J Rice / Anchi Cheng / Carl J Negro / Lawrence Shapiro / Peter D Kwong / David Jeruzalmi / Amedee des Georges / Clinton S Potter / Bridget Carragher /
Abstract: Single particle cryo-electron microscopy (cryoEM) is often performed under the assumption that particles are not adsorbed to the air-water interfaces and in thin, vitreous ice. In this study, we ...Single particle cryo-electron microscopy (cryoEM) is often performed under the assumption that particles are not adsorbed to the air-water interfaces and in thin, vitreous ice. In this study, we performed fiducial-less tomography on over 50 different cryoEM grid/sample preparations to determine the particle distribution within the ice and the overall geometry of the ice in grid holes. Surprisingly, by studying particles in holes in 3D from over 1000 tomograms, we have determined that the vast majority of particles (approximately 90%) are adsorbed to an air-water interface. The implications of this observation are wide-ranging, with potential ramifications regarding protein denaturation, conformational change, and preferred orientation. We also show that fiducial-less cryo-electron tomography on single particle grids may be used to determine ice thickness, optimal single particle collection areas and strategies, particle heterogeneity, and de novo models for template picking and single particle alignment.
History
DepositionDec 5, 2017-
Header (metadata) releaseDec 20, 2017-
Map releaseDec 20, 2017-
UpdateJan 29, 2020-
Current statusJan 29, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Solid view (volume rendering)
  • Imaged by UCSF Chimera
  • Download
  • Solid view (volume rendering)
  • Imaged by UCSF Chimera
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Movie viewer
Supplemental images

emd_7141.png

Downloads & links

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Map

FileDownload / File: emd_7141.map.gz / Format: CCP4 / Size: 1.9 GB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationTomogram of GDH single particle
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1 Å/pix.
x 570 pix.
= 570. Å		1 Å/pix.
x 958 pix.
= 958. Å		1 Å/pix.
x 926 pix.
= 926. Å

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

generated in cubic-lattice coordinate

Voxel sizeX=Y=Z: 1 Å
Density

Histogram

Histogram (log scale)
Minimum - Maximum-0.04832646 - 0.055116534
Average (Standard dev.)0.011135806 (±0.0041482416)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin00-570
Dimensions958926570
Spacing926958570
CellA: 926.0 Å / B: 958.0 Å / C: 570.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z111
M x/y/z926958570
origin x/y/z0.0000.0000.000
length x/y/z926.000958.000570.000
α/β/γ90.00090.00090.000
start NX/NY/NZ-38-19-20
NX/NY/NZ858082
MAP C/R/S123
start NC/NR/NS00-570
NC/NR/NS926958570
D min/max/mean-0.0480.0550.011

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Supplemental data

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Sample components

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Entire : Glutamate dehydrogenase

EntireName: Glutamate dehydrogenase
Components
  • Complex: Glutamate dehydrogenase

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Supramolecule #1: Glutamate dehydrogenase

SupramoleculeName: Glutamate dehydrogenase / type: complex / ID: 1 / Parent: 0 / Details: Tomography on single particle sample
Source (natural)Organism: unidentified (others)

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Experimental details

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Structure determination

Methodcryo EM
Processingelectron tomography
Aggregation stateparticle

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Sample preparation

VitrificationCryogen name: ETHANE
SectioningOther: NO SECTIONING

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Average electron dose: 2.0 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
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

DetailsAppion-Protomo fiducial-less tilt-series alignment
Final reconstructionAlgorithm: SIMULTANEOUS ITERATIVE (SIRT) / Software - Name: TOMO3D / Number images used: 40

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