Summary
Chloride channels were detergent-extracted fromTorpedo electroplax plasma membrane vesicles and reconstituted into liposomes by rapid detergent removal and a freeze-thawsonication procedure. Concentrative uptake of36Cl−, driven by a Cl− gradient was used to determine conductance properties of reconstituted channels. Chloride flux assayed by this method is strongly selective for Cl− over cations, is blocked by SCN−, inactivated by treatment with DIDS, and exhibits an anion selectivity sequence Cl−>Br−>F−>SO 2−4 , as does the voltagegated Cl− channel fromTorpedo observed in planar lipid bilayers. The channels are localized to the noninnervated face of the electrocyte, and a novel trapped-volume method is used to estimate a channel density on the order of 500 pmol/mg protein. An initial fractionation of the membrane extract by anion exchange chromatography yields fivefold enrichment of the channel activity.
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Goldberg, A.F.X., Miller, C. Solubilization and functional reconstitution of a chloride channel fromTorpedo californica electroplax. J. Membrain Biol. 124, 199–206 (1991). https://doi.org/10.1007/BF01994354
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DOI: https://doi.org/10.1007/BF01994354