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- EMDB-1618: 3D reconstruction of heterodimeric yeast Pol alpha using electron... -

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Basic information

Entry
Database: EMDB / ID: 1618
Title3D reconstruction of heterodimeric yeast Pol alpha using electron microscopy
KeywordsDNA replication / DNA Polymerase alpha / Electron microscopy
SampleYeast DNA polymerase alpha
SourceSaccharomyces cerevisiae / yeast / Baker's yeast / サッカロミセス・セレビシエ /
Map data3D reconstruction of heterodimeric yeast Pol alpha using electron microscopy
Methodsingle particle reconstruction, at 22.9 Å resolution
AuthorsKlinge S / Nunez-Ramirez R / Llorca O / Pellegrini L
CitationEMBO J., 2009, 28, 1978-1987

EMBO J., 2009, 28, 1978-1987 Yorodumi Papers
3D architecture of DNA Pol alpha reveals the functional core of multi-subunit replicative polymerases.
Sebastian Klinge / Rafael Núñez-Ramírez / Oscar Llorca / Luca Pellegrini

DateDeposition: May 4, 2009 / Header (metadata) release: May 7, 2009 / Map release: May 6, 2011 / Last update: May 6, 2011

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.045
  • Imaged by UCSF CHIMERA
  • Download
  • Surface view colored by radius
  • Surface level: 0.045
  • Imaged by UCSF CHIMERA
  • Download
3D viewer


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Supplemental images

Downloads & links

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Map

Fileemd_1618.map.gz (map file in CCP4 format, 1025 KB)
Projections & slices

Image control

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AxesZ (Sec.)Y (Row.)X (Col.)
64 pix
3.8 Å/pix.
= 243.2 Å
64 pix
3.8 Å/pix.
= 243.2 Å
64 pix
3.8 Å/pix.
= 243.2 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider package.

Voxel sizeX=Y=Z: 3.8 Å
Density
Contour Level:0.045 (by author), 0.045 (movie #1):
Minimum - Maximum-0.0982105 - 0.13849
Average (Standard dev.)0.00211433 (0.0123166)
Details

EMDB XML:

Space Group Number1
Map Geometry
Axis orderXYZ
Dimensions646464
Origin000
Limit636363
Spacing646464
CellA=B=C: 243.2 Å
α=β=γ: 90 deg.

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z3.83.83.8
M x/y/z646464
origin x/y/z-0.000-0.000-0.000
length x/y/z243.200243.200243.200
α/β/γ90.00090.00090.000
start NX/NY/NZ-17-17-200
NX/NY/NZ123123401
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS646464
D min/max/mean-0.0980.1380.002

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Supplemental data

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Sample components

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Entire Yeast DNA polymerase alpha

EntireName: Yeast DNA polymerase alpha / Number of components: 2
Oligomeric State: One monomer of DNA polymerase alpha binds to a monomer of B-subunit
MassTheoretical: 185 kDa / Measured by: Gel Filtration

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Component #1: protein, DNA polymerase alpha

ProteinName: DNA polymerase alpha / a.k.a: DNA polymerase alpha / Oligomeric Details: Dimer / Number of Copies: 1 / Recombinant expression: Yes
MassTheoretical: 130 kDa / Experimental: 130 kDa
SourceSpecies: Saccharomyces cerevisiae / yeast / Baker's yeast / サッカロミセス・セレビシエ /
External referencesGene Ontology: GO: 0005658, GO: 0005739, GO: 0003677, GO: 0003887, GO: 0001882, GO: 0000166, GO: 0005515, GO: 0006270, GO: 0000731, GO: 0006273, GO: 0006279, GO: 0006278
InterPro: InterPro: 006172, InterPro: 006134, InterPro: 017966, InterPro: 017964, InterPro: 006133, InterPro: 004578, InterPro: 015088

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Component #2: protein, B subunit

ProteinName: B subunit / a.k.a: B subunit / Oligomeric Details: Dimer / Recombinant expression: Yes / Number of Copies: 1
MassTheoretical: 55 kDa / Experimental: 55 kDa
SourceSpecies: Saccharomyces cerevisiae / yeast / Baker's yeast / サッカロミセス・セレビシエ /
External referencesInterPro: InterPro: 007185, InterPro: 016722, InterPro: 013627
Gene Ontology: GO: 0005658, GO: 0005635, GO: 0003677, GO: 0003887, GO: 0005515, GO: 0006270, GO: 0006273, GO: 0016233

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Experimental details

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Sample preparation

Specimen stateparticle
Sample solutionSpecimen conc.: 0.7 mg/ml / Buffer solution: 100mM TrisHCl pH 8.0, 500 mM NaCl / pH: 8
Support film400 mesh copper grid
StainingA diluted solution of Pol alpha complex was adsorbed onto glow-discharged carbon coated grids, stained with 2% uranyl formate
VitrificationInstrument: NONE / Cryogen name: NONE

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Electron microscopy imaging

ImagingMicroscope: JEOL 1230 / Date: Aug 20, 2009 / Details: Microscope used JEOL-1230
Electron gunElectron source: TUNGSTEN HAIRPIN / Accelerating voltage: 100 kV / Electron dose: 15 e/Å2 / Illumination mode: FLOOD BEAM
LensMagnification: 50000 X (nominal)
Astigmatism: Objective lens astigmatism was corrected at 120,000 times magnification
Cs: 2.9 mm / Imaging mode: BRIGHT FIELD / Defocus: 750 - 1200 nm
Specimen HolderHolder: Eucentric / Model: JEOL / Temperature: 293 K ( 293 - 293 K)
CameraDetector: KODAK SO-163 FILM

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Image acquisition

Image acquisitionNumber of digital images: 170 / Scanner: OTHER / Sampling size: 10 microns / Bit depth: 8

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Image processing

ProcessingMethod: single particle reconstruction / Number of class averages: 190 / Number of projections: 12913 / Applied symmetry: C1 (asymmetric)
3D reconstructionAlgorithm: ML3D / Software: EMAN, Xmipp / CTF correction: reverse phases / Resolution: 22.9 Å / Resolution method: FSC 0.5

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Atomic model buiding

Modeling #1Software: adpem / Refinement protocol: rigid body / Target criteria: cross-correlation / Refinement space: REAL / Details: Protocol: Rigid Body
Input PDB model: 3FLO
Modeling #2Software: adpem / Refinement protocol: rigid body / Target criteria: cross-correlation / Refinement space: REAL / Details: Protocol: Rigid Body
Input PDB model: 2VWJ

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