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L-Type Ca Ion Channel

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L-Type Ca Ion Channel

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Visualization of the domain structure of an L-type Ca2+ channel using electron cryo-microscopy

Abstract

The three-dimensional structure of the skeletal muscle voltage-gated L-type calcium channel (Ca(v)1.1; dihydropyridine receptor, DHPR) was determined using electron cryo-microscopy and single-particle averaging. The structure shows a single channel complex with an approximate total molecular mass of 550 kDa, corresponding to the five known subunits of the DHPR, and bound detergent and lipid. Features visible in our structure together with antibody labeling of the beta and alpha(2) subunits allowed us to assign locations for four of the five subunits within the structure. The most striking feature of the structure is the extra-cellular alpha(2) subunit that protrudes from the membrane domain in close proximity to the alpha(1) subunit. The cytosolic beta subunit is located close to the membrane and adjacent to subunits alpha(1), gamma and delta. Our structure correlates well with the functional and biochemical data available for this channel and suggests a three-dimensional model for the excitation-contraction coupling complex consisting of DHPR tetrads and the calcium release channel.

Data Description

Following the link to the right goes to a stack (952 MB) of 15,229 images (MRC/CCP4 format) of L-type Calcium channel (DHPR [1]). The images were recorded with a FEI Tecnai F20 electron cryo-mircroscope equipped with a field emission gun operating at 200 kV. Micrographs were taken under low-dose conditions at an underfocus between 3.0 and 4.5 µm and magnification of 62,000x, which were then compressed threefold resulting in a pixel size of 3.39 Å/pixel at the specimen level. Alignment parameters determined by Frealign for each particle in the stack can also be found below, as well as a reconstruction of DHPR (also MRC/CCP4 format; a B-factor of -3000.0 Å2 and 23.0 Å low-pass filter radius were applied using the program bfactor to sharpen the density) and a script to calculate a reconstruction using Frealign.

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