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Crystal structure and molecular imaging of the Nav channel β3 subunit indicates a trimeric assembly.

Accepted version
Peer-reviewed

Type

Article

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Authors

Namadurai, Sivakumar 
Balasuriya, Dilshan 
Rajappa, Rajit 
Wiemhöfer, Martin 

Abstract

The vertebrate sodium (Nav) channel is composed of an ion-conducting α subunit and associated β subunits. Here, we report the crystal structure of the human β3 subunit immunoglobulin (Ig) domain, a functionally important component of Nav channels in neurons and cardiomyocytes. Surprisingly, we found that the β3 subunit Ig domain assembles as a trimer in the crystal asymmetric unit. Analytical ultracentrifugation confirmed the presence of Ig domain monomers, dimers, and trimers in free solution, and atomic force microscopy imaging also detected full-length β3 subunit monomers, dimers, and trimers. Mutation of a cysteine residue critical for maintaining the trimer interface destabilized both dimers and trimers. Using fluorescence photoactivated localization microscopy, we detected full-length β3 subunit trimers on the plasma membrane of transfected HEK293 cells. We further show that β3 subunits can bind to more than one site on the Nav 1.5 α subunit and induce the formation of α subunit oligomers, including trimers. Our results suggest a new and unexpected role for the β3 subunits in Nav channel cross-linking and provide new structural insights into some pathological Nav channel mutations.

Description

Keywords

Analytical Ultracentrifugation, Atomic Force Microscopy, In Vivo Imaging, Photoactivated Localization Microscopy, Protein Trimerization, Sodium Channels, X-ray Crystallography, Amino Acid Sequence, Binding Sites, Cloning, Molecular, Crystallization, Crystallography, X-Ray, Dimerization, HEK293 Cells, Humans, Immunoglobulins, Microscopy, Atomic Force, Molecular Sequence Data, NAV1.5 Voltage-Gated Sodium Channel, Protein Conformation, Ultracentrifugation, Voltage-Gated Sodium Channel beta-3 Subunit

Journal Title

J Biol Chem

Conference Name

Journal ISSN

0021-9258
1083-351X

Volume Title

289

Publisher

Elsevier BV
Sponsorship
Wellcome Trust (089125/Z/09/Z)