Journal Article

Three-dimensional structure of the α<sub>1</sub>–β complex in the skeletal muscle dihydropyridine receptor by single-particle electron microscopy

Kazuyoshi Murata, Shinpei Nishimura, Akihiko Kuniyasu and Hitoshi Nakayama

in Microscopy

Published on behalf of The Japanese Society of Microscopy

Volume 59, issue 3, pages 215-226
Published in print June 2010 | ISSN: 2050-5698
Published online December 2009 | e-ISSN: 2050-5701 | DOI: https://dx.doi.org/10.1093/jmicro/dfp059
Three-dimensional structure of the α1–β complex in the skeletal muscle dihydropyridine receptor by single-particle electron microscopy

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The dihydropyridine receptor (DHPR) is a protein complex that consists of five distinct subunits of α1, α2, β, γ and δ and functions as a voltage-dependent L-type Ca2+ channel. Here we purified the α1–β complex (∼250 kDa) from the rabbit skeletal muscle DHPR and reconstructed its three-dimensional (3D) structure to 38 Å resolution by single particle analysis of negative staining electron microscopy. The α1–β structure exhibited two unique regions: a pseudo-4-fold petaloid region and an elongated region. X-ray crystallographic models of a homologous voltage-dependent K+ channel and the β subunit fit well into the individual regions of the α1–β structure, revealing that the two regions correspond to the transmembrane α1 and the cytoplasmic β subunits, respectively. In addition, 3D reconstruction and immuno-electron microscopic analysis performed on the independently purified DHPR demonstrated that the α1–β complex was located in the large globular portion of the DHPR, and the N-terminal region of the β subunit was extended to the leg-shaped protrusion of the DHPR, which includes the α2δ subunits. Our results propose a model in which the β subunit may regulate ion channel function by acting as a hinge between α1 and α2δ subunits of the DHPR.

Keywords: membrane protein; voltage-dependent ion channel; skeletal muscle; electron microscopy; single particle analysis; negative staining

Journal Article.  6776 words.  Illustrated.

Subjects: Biological Sciences

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