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bioRxiv · 10.1101/2024.05.20.594950

Isoform-specific N-linked glycosylation of voltage-gated sodium channel alpha-subunits alters beta-subunit binding sites

Beaudoin, C. A.·Kohli, M.·Salvage, S. C.·Liu, H.·Arundel, S. J.·Hamaia, S. W.·Lei, M.·Huang, C. L.- H.·Jackson, A. P.
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Abstract

HighlightsO_LIThree N-linked glycosylation sites conserved among all Nav channels C_LIO_LIGlycan modelling and molecular dynamics simulations highlight 3D landscape C_LIO_LIUnique Nav1.5 N-linked glycans may prevent binding to Ig-domains of {beta}1 and {beta}3 C_LIO_LIUnique Nav1.8 N-linked glycan may prevent binding to Ig-domains of {beta}2 and {beta}4 C_LIO_LIN-linked glycans likely contribute to supra-molecular clustering of Nav channels C_LI Voltage-gated sodium channel -subunits (Nav1.1-1.9) initiate and propagate action potentials in neurons and myocytes. The Nav {beta}-subunits ({beta}1-4) have been shown to modulate -subunit properties. Homo-oligomerization of {beta}-subunits on neighboring or opposing plasma membranes has been suggested to facilitate cis or trans interactions, respectively. The interactions between several Nav channel isoforms and {beta}-subunits have been determined using cryogenic electron microscopy (cryo-EM). Interestingly, the Nav cryo-EM structures reveal the presence of N-linked glycosylation sites. However, only the first glycan moieties are typically resolved at each site due to the flexibility of mature glycan trees. Thus, existing cryo-EM structures may risk de-emphasizing the structural implications of glycans on the Nav channels. Herein, molecular modelling and all-atom molecular dynamics simulations were applied to investigate the conformational landscape of N-linked glycans on Nav channel surfaces. The simulations revealed that negatively-charged sialic acid residues of two glycan sites may interact with voltage-sensing domains. Notably, two Nav1.5 isoform-specific glycans extensively cover the -subunit region that, in other Nav channel -subunit isoforms, corresponds to the binding site for the {beta}1-(and likely {beta}3-) subunit immunoglobulin (Ig) domain. Nav1.8 contains a unique N-linked glycosylation site that likely prevents its interaction with the {beta}2 and {beta}4-subunit Ig domain. These isoform-specific glycans may have evolved to facilitate specific functional interactions, for example by redirecting {beta}-subunit Ig-domains outwards to permit cis or trans supra-clustering within specialized cellular compartments such as the cardiomyocyte perinexal space. Further experimental work is necessary to validate these predictions.

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Beaudoin, C. A., Kohli, M., Salvage, S. C., Liu, H., Arundel, S. J., Hamaia, S. W., Lei, M., Huang, C. L.- H., Jackson, A. P.. 2024-05-20. Isoform-specific N-linked glycosylation of voltage-gated sodium channel alpha-subunits alters beta-subunit binding sites. https://doi.org/10.1101/2024.05.20.594950

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