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Yorodumi- PDB-8emq: Mouse apoferritin heavy chain with zinc determined using single-p... -
+Open data
-Basic information
Entry | Database: PDB / ID: 8emq | ||||||||||||
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Title | Mouse apoferritin heavy chain with zinc determined using single-particle cryo-EM with Apollo camera. | ||||||||||||
Components | Ferritin heavy chain, N-terminally processed | ||||||||||||
Keywords | METAL BINDING PROTEIN / bind with iron | ||||||||||||
Function / homology | Function and homology information Iron uptake and transport / Golgi Associated Vesicle Biogenesis / iron ion sequestering activity / negative regulation of ferroptosis / ferroxidase / autolysosome / intracellular sequestering of iron ion / ferroxidase activity / negative regulation of fibroblast proliferation / endocytic vesicle lumen ...Iron uptake and transport / Golgi Associated Vesicle Biogenesis / iron ion sequestering activity / negative regulation of ferroptosis / ferroxidase / autolysosome / intracellular sequestering of iron ion / ferroxidase activity / negative regulation of fibroblast proliferation / endocytic vesicle lumen / autophagosome / Neutrophil degranulation / ferric iron binding / ferrous iron binding / iron ion transport / immune response / iron ion binding / negative regulation of cell population proliferation / mitochondrion / extracellular region / identical protein binding / membrane / cytosol / cytoplasm Similarity search - Function | ||||||||||||
Biological species | Mus musculus (house mouse) | ||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 1.66 Å | ||||||||||||
Authors | Peng, R. / Fu, X. / Mendez, J.H. / Randolph, P.H. / Bammes, B. / Stagg, S.M. | ||||||||||||
Funding support | United States, 3items
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Citation | Journal: J Struct Biol X / Year: 2023 Title: Characterizing the resolution and throughput of the Apollo direct electron detector. Authors: Ruizhi Peng / Xiaofeng Fu / Joshua H Mendez / Peter S Randolph / Benjamin E Bammes / Scott M Stagg / Abstract: Advances in electron detection have been essential to the success of high-resolution cryo-EM structure determination. A new generation of direct electron detector called the Apollo, has been ...Advances in electron detection have been essential to the success of high-resolution cryo-EM structure determination. A new generation of direct electron detector called the Apollo, has been developed by Direct Electron. The Apollo uses a novel event-based MAPS detector custom designed for ultra-fast electron counting. We have evaluated this new camera, finding that it delivers high detective quantum efficiency (DQE) and low coincidence loss, enabling high-quality electron counting data acquisition at up to nearly 80 input electrons per pixel per second. We further characterized the performance of Apollo for single particle cryo-EM on real biological samples. Using mouse apoferritin, Apollo yielded better than 1.9 Å resolution reconstructions at all three tested dose rates from a half-day data collection session each. With longer collection time and improved specimen preparation, mouse apoferritin was reconstructed to 1.66 Å resolution. Applied to a more challenging small protein aldolase, we obtained a 2.24 Å resolution reconstruction. The high quality of the map indicates that the Apollo has sufficiently high DQE to reconstruct smaller proteins and complexes with high-fidelity. Our results demonstrate that the Apollo camera performs well across a broad range of dose rates and is capable of capturing high quality data that produce high-resolution reconstructions for large and small single particle samples. | ||||||||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8emq.cif.gz | 1.7 MB | Display | PDBx/mmCIF format |
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PDB format | pdb8emq.ent.gz | 1.4 MB | Display | PDB format |
PDBx/mmJSON format | 8emq.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 8emq_validation.pdf.gz | 1 MB | Display | wwPDB validaton report |
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Full document | 8emq_full_validation.pdf.gz | 1.1 MB | Display | |
Data in XML | 8emq_validation.xml.gz | 119.4 KB | Display | |
Data in CIF | 8emq_validation.cif.gz | 157.3 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/em/8emq ftp://data.pdbj.org/pub/pdb/validation_reports/em/8emq | HTTPS FTP |
-Related structure data
Related structure data | 28259MC 8ehgC 8en7C M: map data used to model this data C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
#1: Protein | Mass: 20079.594 Da / Num. of mol.: 24 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Mus musculus (house mouse) / Gene: Fth1, Fth / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P09528 #2: Chemical | ChemComp-ZN / #3: Chemical | ChemComp-FE / #4: Water | ChemComp-HOH / | Has ligand of interest | Y | |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: Mouse apoferritin heavy chain / Type: COMPLEX Details: Ferritin is a universal intracellular protein that stores iron and releases it in a controlled fashion. Entity ID: #1 / Source: RECOMBINANT | ||||||||||||||||||||
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Molecular weight | Value: 0.506 MDa / Experimental value: NO | ||||||||||||||||||||
Source (natural) | Organism: Mus musculus (house mouse) | ||||||||||||||||||||
Source (recombinant) | Organism: Escherichia coli BL21(DE3) (bacteria) | ||||||||||||||||||||
Buffer solution | pH: 7.5 / Details: DTT are added freshly before use. | ||||||||||||||||||||
Buffer component |
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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: UltrAuFoil R1.2/1.3 | ||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 95 % / Chamber temperature: 277.15 K |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 75000 X / Calibrated magnification: 72621 X / Nominal defocus max: 1500 nm / Nominal defocus min: 400 nm / Cs: 2.7 mm / C2 aperture diameter: 80 µm / Alignment procedure: ZEMLIN TABLEAU |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 1.597 sec. / Electron dose: 60 e/Å2 / Film or detector model: DIRECT ELECTRON APOLLO (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 3804 |
Image scans | Sampling size: 4.34 µm / Width: 8192 / Height: 8192 |
-Processing
Software |
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EM software |
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Image processing | Details: Direct Electron Apollo | |||||||||||||||||||||||||||||||||||
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | |||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 183073 | |||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 1.66 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 286970 / Symmetry type: POINT | |||||||||||||||||||||||||||||||||||
Refinement | Cross valid method: NONE |