Detergent-free solubilization of human Kv channels expressed in mammalian cellsстатья

Информация о цитировании статьи получена из Scopus, Web of Science
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Дата последнего поиска статьи во внешних источниках: 25 апреля 2019 г.

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[1] Detergent-free solubilization of human kv channels expressed in mammalian cells / M. G. Karlova⁠, N. Voskoboynikova⁠, G. S. Gluhov⁠ et al. // Chemistry and Physics of Lipids. — 2019. — Vol. 219. — P. 50–57. Styrene-maleic acid (SMA) copolymers are used to extract lipid-encased membrane proteins from lipid bilayers in a detergent-free manner, yielding SMA lipid particles (SMALPs). SMALPs can serve as stable water-soluble nanocontainers for structural and functional studies of membrane proteins. Here, we used SMA copolymers to study full-length pore-forming α-subunits hKCNH5 and hKCNQ1 of human neuronal and cardiac voltage-gated potassium (Kv) channels, as well as the fusion construct comprising of an α-subunit hKCNQ1 and its regulatory transmembrane KCNE1 β-subunit (hKCNE1-hKCNQ1) with added affinity tags, expressed in mammalian COS-1 cells. All these recombinant proteins were shown to be functionally active. Treatment with the SMA copolymer, followed by purification on the affinity column, enabled extraction of all three channels. A DLS experiment demonstrated that negative stain electron microscopy and single particle image analysis revealed a four-fold symmetry within channel-containing SMALPs, which indicates that purified hKCNH5 and hKCNQ1 channels, as well as the hKCNE1-hKCNQ1 fusion construct, retained their structural integrity as tetramers. [ DOI ]

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