REL PDBe PDBe PDBe 2004-03-10 2004-03-11 2004-03-11 2012-10-24 Visualization of the domain structure of an L-type Ca2+ channel using electron cryo-microscopy. Wolf M Eberhart A Glossmann H Striessnig J Grigorieff N Wolf M Eberhart A Glossmann H Striessnig J Grigorieff N Visualization of the domain structure of an L-type Ca2+ channel using electron cryo-microscopy. J.MOL.BIOL. 332 171 182 2003 12946355 doi:10.1016/S0022-2836(03)00899-4 skeletal muscle dihydropyridine receptor skeletal muscle dihydropyridine receptor
channel complex is solubilized in Digitonin
monodisperse channel-detergent complex consisting of 5 subunits 1 0.8 0.55 gel filtration chromatography DHPR with subunits alpha1, alpha2, beta, gamma, delta Oryctolagus cuniculus Rabbit rabbit skeletal muscle animal tissue sarcoplasmatic reticulum t tubular junction 0.55 0.8
additional MW due to detergent and residual lipid; solubilized in Digitonin
1 monodisperse channel detergent complex with 5 subunits false singleParticle particle 0.35 7.4
50mM Tris-HCl pH7.4, 150mM NaCl, 25uM CaCl2, 1mM Iodoacetamide, 0.1mM Benzamidine, 0.1mM PMSF, 0.1% (w/v) Digitonin.
 NEGATIVE
5ul sample on holey carbon grids (Quantifoil) were vitrified by plunging into liqid ethane after blotting with filter paper.
Quantifoil (R) holey carbon copper grids
 ETHANE 80 20 HOMEMADE PLUNGER
Vitrification instrument: custom plunger
Blot 5ul sample on negative glow-discharged grids for 5-10sec with Whatman filter paper before plunging. FEI TECNAI F20 FLOOD BEAM BRIGHT FIELD FIELD EMISSION GUN 200 2.0 3.0 4.5 62000.0 GATAN LIQUID NITROGEN 90 95 90 obj lens astigmatism was corrected at 270,000x magnification 2002-02-05 KODAK SO-163 FILM ZEISS SCAI 7 180 9
full scan resolution was downsampled by 3x3 pixel averaging.
1.0 8. Side entry liquid nitrogen-cooled cryo holder 0 0
per micrograph, CTFFIT3
 C1 OTHER 21.0 FSC 0.5 CUT-OFF MRC, IMAGIC, FREALIGN
FREALIGN does not use class averages but performs an orientation search of each individual particle incl. CTF correction. The final map was reconstructed from contributions of 14,056 particles and sharpened with a negative temperature factor of 500. The absolute handedness of the reconstruction was not determined.
14056 emd_1069.map.gz 1 IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) 128 128 128 -64 -64 -64 128 128 128 433.92 433.92 433.92 90 90 90 X Y Z -0.0164905 0.0980307 0.0013486 0.00896609 3.39 3.39 3.39 0.0148 3D reconstruction of the skeletal muscle dihydropyridin receptor (DHPR, L-type calcium channel) from single particle images. Low-pass filtered at 23A resolution. Ref.: J Mol Biol. 2003 Sep 5;332(1):171-82 If displayed with chimera (http://www.cgl.ucsf.edu/chimera/download) threshold level for published protein surface (white transparent) = 0.0626 threshold level for published core density (red) = 0.0763
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