Abstract
The L-type Ca2+ channel (dihydropyridine receptor (DHPR) in skeletal muscle acts as the voltage sensor for excitation-contraction coupling. To better resolve the spatial organization of the DHPR subunits (α1s or CaV1.1, α2, β1a, δ1, and γ), we created transgenic mice expressing a recombinant β1a subunit with YFP and a biotin acceptor domain attached to its N- and C- termini, respectively. DHPR complexes were purified from skeletal muscle, negatively stained, imaged by electron microscopy, and subjected to single-particle image analysis. The resulting 19.1-Åresolution, three-dimensional reconstruction shows a main body of 17 x 11 x 8 nm with five corners along its perimeter. Two protrusions emerge from either face of the main body: the larger one attributed to the α2-δ1 subunit that forms a flexible hook-shaped feature and a smaller protrusion on the opposite side that corresponds to the II-III loop of CaV1.1 as revealed by antibody labeling. Novel features discernible in the electron density accommodate the atomic coordinates of a voltage-gated sodium channel and of the β subunit in a single docking possibility that defines the α1-β interaction. The β subunit appears more closely associated to the membrane than expected, which may better account for both its role in localizing the α1s subunit to the membrane and its suggested role in excitation-contraction coupling.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 43853-43861 |
| Number of pages | 9 |
| Journal | Journal of Biological Chemistry |
| Volume | 287 |
| Issue number | 52 |
| DOIs | |
| State | Published - Dec 21 2012 |
| Externally published | Yes |
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ASJC Scopus subject areas
- Biochemistry
- Cell Biology
- Molecular Biology
Cite this
Three-dimensional localization of the α and β subunits and of the II-III loop in the skeletal muscle L-type Ca2+ channel. / Szpyt, John; Lorenzon, Nancy; Perez, Claudio F.; Norris, Ethan; Allen, Paul D.; Beam, Kurt G.; Samsó, Montserrat.
In: Journal of Biological Chemistry, Vol. 287, No. 52, 21.12.2012, p. 43853-43861.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Three-dimensional localization of the α and β subunits and of the II-III loop in the skeletal muscle L-type Ca2+ channel
AU - Szpyt, John
AU - Lorenzon, Nancy
AU - Perez, Claudio F.
AU - Norris, Ethan
AU - Allen, Paul D.
AU - Beam, Kurt G.
AU - Samsó, Montserrat
PY - 2012/12/21
Y1 - 2012/12/21
N2 - The L-type Ca2+ channel (dihydropyridine receptor (DHPR) in skeletal muscle acts as the voltage sensor for excitation-contraction coupling. To better resolve the spatial organization of the DHPR subunits (α1s or CaV1.1, α2, β1a, δ1, and γ), we created transgenic mice expressing a recombinant β1a subunit with YFP and a biotin acceptor domain attached to its N- and C- termini, respectively. DHPR complexes were purified from skeletal muscle, negatively stained, imaged by electron microscopy, and subjected to single-particle image analysis. The resulting 19.1-Åresolution, three-dimensional reconstruction shows a main body of 17 x 11 x 8 nm with five corners along its perimeter. Two protrusions emerge from either face of the main body: the larger one attributed to the α2-δ1 subunit that forms a flexible hook-shaped feature and a smaller protrusion on the opposite side that corresponds to the II-III loop of CaV1.1 as revealed by antibody labeling. Novel features discernible in the electron density accommodate the atomic coordinates of a voltage-gated sodium channel and of the β subunit in a single docking possibility that defines the α1-β interaction. The β subunit appears more closely associated to the membrane than expected, which may better account for both its role in localizing the α1s subunit to the membrane and its suggested role in excitation-contraction coupling.
AB - The L-type Ca2+ channel (dihydropyridine receptor (DHPR) in skeletal muscle acts as the voltage sensor for excitation-contraction coupling. To better resolve the spatial organization of the DHPR subunits (α1s or CaV1.1, α2, β1a, δ1, and γ), we created transgenic mice expressing a recombinant β1a subunit with YFP and a biotin acceptor domain attached to its N- and C- termini, respectively. DHPR complexes were purified from skeletal muscle, negatively stained, imaged by electron microscopy, and subjected to single-particle image analysis. The resulting 19.1-Åresolution, three-dimensional reconstruction shows a main body of 17 x 11 x 8 nm with five corners along its perimeter. Two protrusions emerge from either face of the main body: the larger one attributed to the α2-δ1 subunit that forms a flexible hook-shaped feature and a smaller protrusion on the opposite side that corresponds to the II-III loop of CaV1.1 as revealed by antibody labeling. Novel features discernible in the electron density accommodate the atomic coordinates of a voltage-gated sodium channel and of the β subunit in a single docking possibility that defines the α1-β interaction. The β subunit appears more closely associated to the membrane than expected, which may better account for both its role in localizing the α1s subunit to the membrane and its suggested role in excitation-contraction coupling.
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UR - http://www.scopus.com/inward/citedby.url?scp=84871532373&partnerID=8YFLogxK
U2 - 10.1074/jbc.M112.419283
DO - 10.1074/jbc.M112.419283
M3 - Article
C2 - 23118233
AN - SCOPUS:84871532373
VL - 287
SP - 43853
EP - 43861
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
SN - 0021-9258
IS - 52
ER -