Preparation of vesicles containing natural and synthetic ion channels

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Jun Zhang1, Rebecca B. Price2, Peter A.V. Anderson2, Kun Fang1, Christopher R. Williams2, Frank J. Raucci3, Joanna R. Long3, Arthur S. Edison3, and Randolph S. Duran1. (1) Butler Polymer Laboratory, Department of Chemistry, University of Florida, Gainesville, FL 36211, (2) Whitney Laboratory, University of Florida, St. Augustine, FL 32080, (3) Biochemistry and Molecular Biology, University of Florida, Gainesville, FL 36211
Large conductance calcium-activated potassium (Maxi-K) ion channels derived from the Slowpoke (Slo) gene could be cloned and heterogeneously expressed in Xenopus oocytes. Vesicles containing cloned and expressed Maxi-K channels from Xenopus oocyte membranes represent some well-defined model systems for electrophysiological investigations of channel structure and function. In this report, the preparation of vesicles under different conditions was studied. Both large (~20 µm) and small unilamellar (~2µm) vesicles were obtained. Large vesicles can be used to study functional properties characteristic of native Maxi-K channels by patch-clamp directly, while unilamellar vesicles are very important in the reconstitution of ion channels into lipid bilayers tethered over semiconductor devices. M2, the transmembrane sequence of the Acetylcholine Receptor and the FMRFamide-gated sodium channel were also investigated.