+
Open data
-
Basic information
Entry | Database: PDB / ID: 8cpu | ||||||
---|---|---|---|---|---|---|---|
Title | Human apoferritin after 561 nm laser exposure | ||||||
![]() | Ferritin heavy chain, N-terminally processed | ||||||
![]() | METAL BINDING PROTEIN / Iron binding protein | ||||||
Function / homology | ![]() iron ion sequestering activity / : / autolysosome / Scavenging by Class A Receptors / Golgi Associated Vesicle Biogenesis / ferroxidase / intracellular sequestering of iron ion / ferroxidase activity / negative regulation of fibroblast proliferation / ferric iron binding ...iron ion sequestering activity / : / autolysosome / Scavenging by Class A Receptors / Golgi Associated Vesicle Biogenesis / ferroxidase / intracellular sequestering of iron ion / ferroxidase activity / negative regulation of fibroblast proliferation / ferric iron binding / Iron uptake and transport / ferrous iron binding / tertiary granule lumen / iron ion transport / intracellular iron ion homeostasis / ficolin-1-rich granule lumen / iron ion binding / immune response / negative regulation of cell population proliferation / Neutrophil degranulation / extracellular exosome / extracellular region / identical protein binding / membrane / nucleus / cytoplasm / cytosol Similarity search - Function | ||||||
Biological species | ![]() | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 1.76 Å | ||||||
![]() | Last, M.G.F. / Noteborn, W.E.M. / Sharp, T.H. | ||||||
Funding support | ![]()
| ||||||
![]() | ![]() Title: Super-resolution fluorescence imaging of cryosamples does not limit achievable resolution in cryoEM. Authors: Mart G F Last / Willem E M Noteborn / Lenard M Voortman / Thomas H Sharp / ![]() Abstract: Correlated super-resolution cryo-fluorescence and cryo-electron microscopy (cryoEM) has been gaining popularity as a method to investigate biological samples with high resolution and specificity. A ...Correlated super-resolution cryo-fluorescence and cryo-electron microscopy (cryoEM) has been gaining popularity as a method to investigate biological samples with high resolution and specificity. A concern in this combined method (called SR-cryoCLEM), however, is whether and how fluorescence imaging prior to cryoEM acquisition is detrimental to sample integrity. In this report, we investigated the effect of high-dose laser light (405, 488, and 561 nm) irradiation on apoferritin samples prepared for cryoEM with excitation wavelengths commonly used in fluorescence microscopy, and compared these samples to controls that were kept in the dark. We found that laser illumination, of equal duration and intensity as used in cryo-single molecule localization microscopy (cryoSMLM) and in the presence of high concentrations of fluorescent protein, did not affect the achievable resolution in cryoEM, with final reconstructions reaching resolutions of ∼ 1.8 Å regardless of the laser illumination. The finding that super-resolution fluorescence imaging of cryosamples prior to cryoEM data acquisition does not limit the achievable resolution suggests that super-resolution cryo-fluorescence microscopy and in situ structural biology using cryoEM are entirely compatible. | ||||||
History |
|
-
Structure visualization
Structure viewer | Molecule: ![]() ![]() |
---|
-
Downloads & links
-
Download
PDBx/mmCIF format | ![]() | 743.1 KB | Display | ![]() |
---|---|---|---|---|
PDB format | ![]() | 602.6 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Summary document | ![]() | 1.2 MB | Display | ![]() |
---|---|---|---|---|
Full document | ![]() | 1.2 MB | Display | |
Data in XML | ![]() | 101.5 KB | Display | |
Data in CIF | ![]() | 150.9 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 16786MC ![]() 8cpmC ![]() 8cpsC ![]() 8cptC ![]() 8cpvC ![]() 8cpwC ![]() 8cpxC M: map data used to model this data C: citing same article ( |
---|---|
Similar structure data | Similarity search - Function & homology ![]() |
-
Links
-
Assembly
Deposited unit | ![]()
|
---|---|
1 |
|
-
Components
#1: Protein | Mass: 20218.570 Da / Num. of mol.: 24 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() #2: Chemical | ChemComp-NA / #3: Chemical | ChemComp-MG / #4: Water | ChemComp-HOH / | Has ligand of interest | N | |
---|
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
---|---|
EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-
Sample preparation
Component | Name: Apoferritin / Type: ORGANELLE OR CELLULAR COMPONENT / Entity ID: #1 / Source: RECOMBINANT | |||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Molecular weight | Value: 0.484 MDa / Experimental value: NO | |||||||||||||||
Source (natural) | Organism: ![]() | |||||||||||||||
Source (recombinant) | Organism: ![]() ![]() | |||||||||||||||
Buffer solution | pH: 8 | |||||||||||||||
Buffer component |
| |||||||||||||||
Specimen | Conc.: 4 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | |||||||||||||||
Specimen support | Grid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3 | |||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 295 K |
-
Electron microscopy imaging
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
---|---|
Microscopy | Model: TFS KRIOS |
Electron gun | Electron source: ![]() |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 130000 X / Nominal defocus max: 1500 nm / Nominal defocus min: 500 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Electron dose: 50 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) |
EM imaging optics | Energyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV |
-
Processing
Software | Name: UCSF ChimeraX / Version: 1.4/v9 / Classification: model building / URL: https://www.rbvi.ucsf.edu/chimerax/ / Os: Windows / Type: package | ||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
EM software |
| ||||||||||||||||||||||||
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
3D reconstruction | Resolution: 1.76 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 9700 / Symmetry type: POINT | ||||||||||||||||||||||||
Atomic model building | PDB-ID: 6Z6U Accession code: 6Z6U / Source name: PDB / Type: experimental model |