Patch Clamp Electrophysiology for the Study of Bacterial Ion Channels in Giant Spheroplasts of E. coli
Ion channel studies have been focused on ion channels from animal and human cells over many years. Based on the knowledge acquired, predominantly over the last 20 years, a large diversity of ion channels exists in cellular membranes of prokaryotes as well. Paradoxically, most of what is known about the structure of eukaryotic ion channels is based on the structure of bacterial channels. This is largely due to the suitability of bacterial cells for functional and structural studies of biological macromolecules in a laboratory environment. Development of the “giant spheroplast” preparation from E. coli cells was instrumental for functional studies of ion channels in the bacterial cell membrane. Here we describe detailed protocols used for the preparation of giant spheroplasts as well as protocols used for the patch-clamp recording of native or heterologously expressed ion channels in E. coli spheroplast membrane.
Key wordsBacteria Archaea Cephalexin Lysozyme Mechanosensitive channels K+ channels Patch clamp Laplace’s law
We wish to thank Maryrose Constantine as well as Drs. Anna Kloda, Evgeny Petrov, and Andrew Battle for critical reading and constructive comments on a draft of the manuscript. This work was supported by the Australian Research Council, the National Health and Medical Research Council of Australia, and Yamada Science Foundation.
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