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- EMDB-17964: Structure of GAPDH determined by cryoEM at 100 keV -

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Basic information

Entry
Database: EMDB / ID: EMD-17964
TitleStructure of GAPDH determined by cryoEM at 100 keV
Map data
Sample
  • Complex: Glyceraldehyde 3-phosphate dehydrogenase
    • Protein or peptide: Glyceraldehyde-3-phosphate dehydrogenaseGlyceraldehyde 3-phosphate dehydrogenase
  • Ligand: NICOTINAMIDE-ADENINE-DINUCLEOTIDE
KeywordsGAPDH / OXIDOREDUCTASE
Function / homology
Function and homology information


glyceraldehyde-3-phosphate dehydrogenase (phosphorylating) / glyceraldehyde-3-phosphate dehydrogenase (NAD+) (phosphorylating) activity / glycolytic process / glucose metabolic process / NAD binding / NADP binding / identical protein binding / metal ion binding
Similarity search - Function
Glyceraldehyde-3-phosphate dehydrogenase, type I / Glyceraldehyde 3-phosphate dehydrogenase, active site / Glyceraldehyde 3-phosphate dehydrogenase active site. / Glyceraldehyde 3-phosphate dehydrogenase, NAD binding domain / Glyceraldehyde 3-phosphate dehydrogenase, NAD(P) binding domain / Glyceraldehyde 3-phosphate dehydrogenase, catalytic domain / Glyceraldehyde/Erythrose phosphate dehydrogenase family / Glyceraldehyde 3-phosphate dehydrogenase, C-terminal domain / Glyceraldehyde 3-phosphate dehydrogenase, NAD binding domain / NAD(P)-binding domain superfamily
Similarity search - Domain/homology
Glyceraldehyde-3-phosphate dehydrogenase
Similarity search - Component
Biological speciesCryptosporidium parvum (eukaryote)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.9 Å
AuthorsMcMullan G / Naydenova K / Mihaylov D / Peet MJ / Wilson H / Yamashita K / Dickerson JL / Chen S / Cannone G / Lee Y ...McMullan G / Naydenova K / Mihaylov D / Peet MJ / Wilson H / Yamashita K / Dickerson JL / Chen S / Cannone G / Lee Y / Hutchings KA / Gittins O / Sobhy M / Wells T / El-Gomati MM / Dalby J / Meffert M / Schulze-Briese C / Henderson R / Russo CJ
Funding support United Kingdom, 6 items
OrganizationGrant numberCountry
Medical Research Council (MRC, United Kingdom)MC UP 120117 United Kingdom
Medical Research Council (MRC, United Kingdom)MC U105184322 United Kingdom
Wellcome Trust220526/B/20/Z United Kingdom
Engineering and Physical Sciences Research CouncilR122522 United Kingdom
Innovate UK103806 United Kingdom
Biotechnology and Biological Sciences Research Council (BBSRC)BB/T003677/1 United Kingdom
CitationJournal: Proc Natl Acad Sci U S A / Year: 2023
Title: Structure determination by cryoEM at 100 keV.
Authors: Greg McMullan / Katerina Naydenova / Daniel Mihaylov / Keitaro Yamashita / Mathew J Peet / Hugh Wilson / Joshua L Dickerson / Shaoxia Chen / Giuseppe Cannone / Yang Lee / Katherine A ...Authors: Greg McMullan / Katerina Naydenova / Daniel Mihaylov / Keitaro Yamashita / Mathew J Peet / Hugh Wilson / Joshua L Dickerson / Shaoxia Chen / Giuseppe Cannone / Yang Lee / Katherine A Hutchings / Olivia Gittins / Mohamed A Sobhy / Torquil Wells / Mohamed M El-Gomati / Jason Dalby / Matthias Meffert / Clemens Schulze-Briese / Richard Henderson / Christopher J Russo /
Abstract: Electron cryomicroscopy can, in principle, determine the structures of most biological molecules but is currently limited by access, specimen preparation difficulties, and cost. We describe a purpose- ...Electron cryomicroscopy can, in principle, determine the structures of most biological molecules but is currently limited by access, specimen preparation difficulties, and cost. We describe a purpose-built instrument operating at 100 keV-including advances in electron optics, detection, and processing-that makes structure determination fast and simple at a fraction of current costs. The instrument attains its theoretical performance limits, allowing atomic resolution imaging of gold test specimens and biological molecular structure determination in hours. We demonstrate its capabilities by determining the structures of eleven different specimens, ranging in size from 140 kDa to 2 MDa, using a fraction of the data normally required. CryoEM with a microscope designed specifically for high-efficiency, on-the-spot imaging of biological molecules will expand structural biology to a wide range of previously intractable problems.
History
DepositionJul 17, 2023-
Header (metadata) releaseNov 29, 2023-
Map releaseNov 29, 2023-
UpdateDec 6, 2023-
Current statusDec 6, 2023Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_17964.png

Downloads & links

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Map

FileDownload / File: emd_17964.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 0.8415 Å
Density
Contour LevelBy AUTHOR: 0.04
Minimum - Maximum-0.12177872 - 0.21569552
Average (Standard dev.)0.00012889736 (±0.0062382407)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 215.424 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_17964_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_17964_half_map_1.map
Projections & Slices
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Histogram

Histogram (log scale)

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Half map: #2

Fileemd_17964_half_map_2.map
Projections & Slices
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Slices (1/2)
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Histogram

Histogram (log scale)

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Sample components

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Entire : Glyceraldehyde 3-phosphate dehydrogenase

EntireName: Glyceraldehyde 3-phosphate dehydrogenase
Components
  • Complex: Glyceraldehyde 3-phosphate dehydrogenase
    • Protein or peptide: Glyceraldehyde-3-phosphate dehydrogenaseGlyceraldehyde 3-phosphate dehydrogenase
  • Ligand: NICOTINAMIDE-ADENINE-DINUCLEOTIDE

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Supramolecule #1: Glyceraldehyde 3-phosphate dehydrogenase

SupramoleculeName: Glyceraldehyde 3-phosphate dehydrogenase / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Cryptosporidium parvum (eukaryote)

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Macromolecule #1: Glyceraldehyde-3-phosphate dehydrogenase

MacromoleculeName: Glyceraldehyde-3-phosphate dehydrogenase / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
EC number: glyceraldehyde-3-phosphate dehydrogenase (phosphorylating)
Source (natural)Organism: Cryptosporidium parvum (eukaryote)
Molecular weightTheoretical: 38.059484 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MHHHHHHSSG RENLYFQGTL GINGFGRIGR LVLRACMERN DITVVAINDP FMDVEYMAYL LKYDSVHGNF NGTVEVSGKD LCINGKVVK VFQAKDPAEI PWGASGAQIV CESTGVFTTE EKASLHLKGG AKKVIISAPP KDNVPMYVMG VNNTEYDPSK F NVISNASC ...String:
MHHHHHHSSG RENLYFQGTL GINGFGRIGR LVLRACMERN DITVVAINDP FMDVEYMAYL LKYDSVHGNF NGTVEVSGKD LCINGKVVK VFQAKDPAEI PWGASGAQIV CESTGVFTTE EKASLHLKGG AKKVIISAPP KDNVPMYVMG VNNTEYDPSK F NVISNASC TTNCLAPLAK IINDKFGIVE GLMTTVHSLT ANQLTVDGPS KGGKDWRAGR CAGNNIIPAS TGAAKAVGKV IP ALNGKLT GMAIRVPTPD VSVVDLTCKL AKPASIEEIY QAVKEASNGP MKGIMGYTSD DVVSTDFIGC KYSSIFDKNA CIA LNDSFV KLISWYDNES GYSNRLVDLA VYVASRGL

UniProtKB: Glyceraldehyde-3-phosphate dehydrogenase

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Macromolecule #2: NICOTINAMIDE-ADENINE-DINUCLEOTIDE

MacromoleculeName: NICOTINAMIDE-ADENINE-DINUCLEOTIDE / type: ligand / ID: 2 / Number of copies: 1 / Formula: NAD
Molecular weightTheoretical: 663.425 Da
Chemical component information

ChemComp-NAD:
NICOTINAMIDE-ADENINE-DINUCLEOTIDE / NAD*YM / Nicotinamide adenine dinucleotide

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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: 7.4
GridModel: UltrAuFoil R0./1 / Material: GOLD
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeJEOL 1400/HR + YPS FEG
Electron beamAcceleration voltage: 100 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.5 µm
Sample stageSpecimen holder model: GATAN 626 SINGLE TILT LIQUID NITROGEN CRYO TRANSFER HOLDER
Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: DECTRIS SINGLA (1k x 1k) / Average electron dose: 40.0 e/Å2

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Image processing

Startup modelType of model: EMDB MAP
EMDB ID:

13256

Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionApplied symmetry - Point group: D2 (2x2 fold dihedral) / Resolution.type: BY AUTHOR / Resolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 19411
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

3cps


Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementSpace: RECIPROCAL
Output model

PDB-8pvf:
Structure of GAPDH determined by cryoEM at 100 keV

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