|Entry||Database: PDB / ID: 5upw|
|Title||CryoEM Structure Refinement by Integrating NMR Chemical Shifts with Molecular Dynamics Simulations|
|Keywords||VIRAL PROTEIN / Cryo-EM / HIV capsid / Chemical shift / Molecular Dynamics / hydrolase / viral protein|
|Specimen source||Human immunodeficiency virus type 1 (NEW YORK-5 ISOLATE) / virus|
|Method||Electron microscopy (5 Å resolution / Helical array / Helical)|
|Citation||J Phys Chem B, 2017, 121, 3853-3863|
SummaryFull reportAbout validation report
|Date||Deposition: Feb 4, 2017 / Release: Mar 1, 2017|
Downloads & links
A: Gag polyprotein
B: Gag polyprotein
C: Gag polyprotein
D: Gag polyprotein
E: Gag polyprotein
F: Gag polyprotein
Mass: 24654.268 Da / Num. of mol.: 6 / Fragment: UNP residues 133-353 / Mutation: A92E
Source: (gene. exp.) Human immunodeficiency virus type 1 (NEW YORK-5 ISOLATE) / virus
References: UniProt: P12493
|Experiment||Method: ELECTRON MICROSCOPY|
|EM experiment||Aggregation state: HELICAL ARRAY / Reconstruction method: HELICAL|
|Component||Name: HIV-1 Capsid Protein Assembly / Type: COMPLEX / Entity ID: 1 / Source: RECOMBINANT|
|Molecular weight||Value: 24 deg. / Units: KILODALTONS/NANOMETER / Experimental value: NO|
|Source (natural)||Organism: Human immunodeficiency virus type 1 (NEW YORK-5 ISOLATE)|
|Source (recombinant)||Organism: Escherichia coli|
|Buffer solution||pH: 8|
|Specimen||Conc.: 2 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES|
|Specimen support||Grid material: COPPER / Grid mesh size: 300 / Grid type: Quantifoil R2/1|
|Vitrification||Instrument: HOMEMADE PLUNGER / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 295 kelvins|
Details: The assembled sample (1.5 microliter) was applied to the carbon side of a glow discharged perforated Quantifoil grid, followed by application of 3 microliter of low salt buffer (100 milimolar NaCl, 50 milimolar Tris pH 8.0) on the back side of the grid, and blotting, from the back side, with a filter paper, before plunge-freezing in liquid ethane
-Electron microscopy imaging
Model: Tecnai Polara / Image courtesy: FEI Company
|Microscopy||Microscope model: FEI POLARA 300|
|Electron gun||Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM|
|Electron lens||Mode: BRIGHT FIELD / Nominal magnification: 31000 / Nominal defocus max: 2200 nm / Nominal defocus min: 700 nm / Cs: 2.2 mm / C2 aperture diameter: 100 mm|
|Specimen holder||Cryogen: NITROGEN / Specimen holder model: SIDE ENTRY, EUCENTRIC|
|Image recording||Average exposure time: 6 sec. / Electron dose: 41 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Number of real images: 523|
|Image scans||Movie frames/image: 30|
|CTF correction||Type: PHASE FLIPPING AND AMPLITUDE CORRECTION|
|Helical symmerty||Angular rotation/subunit: -31.13 deg. / Axial rise/subunit: 6.94 Å / Axial symmetry: C1|
|Particle selection||Number of particles selected: 39712|
|3D reconstruction||Resolution: 5 Å / Resolution method: FSC 0.143 CUT-OFF / Number of particles: 38452 / Algorithm: FOURIER SPACE / Number of class averages: 3 / Symmetry type: HELICAL|
|Atomic model building||Ref protocol: FLEXIBLE FIT / Ref space: REAL|
|Atomic model building||PDB-ID: 4XFX|
Pdb chain residue range: 1-220
-Oct 4, 2017. Three pioneers of this field were awarded Nobel Prize in Chemistry 2017
Three pioneers of this field were awarded Nobel Prize in Chemistry 2017
- Jacques Dubochet (University of Lausanne, Switzerland) is a pioneer of ice-embedding method of EM specimen (as known as cryo-EM), Most of 3DEM structures in EMDB and PDB are obtained using his method.
- Joachim Frank (Columbia University, New York, USA) is a pioneer of single particle reconstruction, which is the most used reconstruction method for 3DEM structures in EMDB and EM entries in PDB. And also, he is a develper of Spider, which is one of the most famous software in this field, and is used for some EM Navigor data (e.g. map projection/slice images).
- Richard Henderson (MRC Laboratory of Molecular Biology, Cambridge, UK) was determined the first biomolecule structure by EM. The first EM entry in PDB, PDB-1brd is determinedby him.
External links: The 2017 Nobel Prize in Chemistry - Press Release
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