top_cornerHomeEMD-2502  Contact EMDataBank 
Image unavailable
Title:Substrate Recruitment Pathways in the Yeast Exosome by Electron Microscopy
Authors:Liu J-J, Bratkowski MA, Liu XQ, Niu C-Y, Ke AL, Wang H-W
Sample:Rrp44-Exosome by Cryo-EM
Method:Single particle reconstruction (24 angstroms resolution)
Red flagLatest update:2014-01-15
Sample
Sample name: Rrp44-Exosome by Cryo-EM
Experimental molecular weight of the sample: 0.350
Components:
ID Type Name Exp. MW (MDa) Theo. MW (MDa) Oligomeric details Recombinant expression Synthetic Organism UniProt identifier GO identifier InterPro identifier Virus identifier Details
1proteinRrp440.11falseSaccharomyces cerevisiae
2proteinRrp430.04falseSaccharomyces cerevisiae
3proteinRrp40.04falseSaccharomyces cerevisiae
4proteinCsl40.03falseSaccharomyces cerevisiae
5proteinRrp450.03falseSaccharomyces cerevisiae
6proteinRrp46-TAP0.03trueSaccharomyces cerevisiae
7proteinRrp410.03falseSaccharomyces cerevisiae
8proteinRrp420.03falseSaccharomyces cerevisiae
9proteinMtr30.03falseSaccharomyces cerevisiae
10proteinRrp400.03falseSaccharomyces cerevisiae
Experiment
Specimen state: Particle
Specimen preparation:
pHSpecimen conc.DetailsStainingSpecimen support details
8.02 mg/mL150mM NaCl, 50mM Tris-HCL,1mM DTT, 2mM MgCl2Quantifoil grids (2/2) with 2~3 nm thin carbon on top
Vitrification:
Cryogen nameHumidityTemp.Instr.MethodTime resolvedDetails
ETHANE100%80 KFEI VITROBOT MARK IV4 ul of the reaction solution was then applied to glow-discharged C-flat grids (1.2/1.3) covered with a layer of continuous carbon with a thickness of ~4nm. The grids were then blotted and plunged into liquid ethane in a FEI Vitrobot Mark IV ms
Imaging:
MicroscopeVoltageIllumination modeImaging modeCsDefocus min.Defocus max.Nominal mag.Calibrated mag.Electron sourceDetectorDetector distanceAstigmatism
FEI TITAN KRIOS120 kVFLOOD BEAMBRIGHT FIELD2.7 mm-500 nm-3500 nm5900059000FIELD EMISSION GUNGENERIC GATAN (4k x 4k) mm

Specimen holderHolder modelTilt min.Tilt max.Energy filterEnergy windowTemp.Temp. min.Temp. max.Beam tiltElectron doseOther detailsDate
FEI TITAN KRIOS AUTOGRID HOLDER°° eV82 K80 K85 K mrad20 e/Å209-JUN-2012
Processing
Software:eman, imagic, relion
CTF correction:both each particle and micrograph
Number of particles:27200
Imposed symmetry:C1
Resolution by author:24 Å
Resolution method:FSC 0.5
Processing details:Micrographs of RE were collected using the AutoEMation software (Lei and Frank, 2005) installed on the microscope at low-dose condition with a dose of ~20 electron/A2 and a defocus value ranging from -1.2 to -4 um. The micrographs were collected on a FEI Eagle CCD camera with a pixel size of 1.5 A.
Scanned images:
Num. imagesSampling sizeOD rangeQuant. bit numberOther detailsScanner
1200 μm/pixel
Fitting:
PDBProtocolTarget crit.SoftwareB valueFitting spacePDB chainDetails
2wp8 rigid bodyChimeraREAL