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Patch Clamp Electrophysiology for the Study of Bacterial Ion Channels in Giant Spheroplasts of E. coli

  • Boris MartinacEmail author
  • Paul R. Rohde
  • Charles G. Cranfield
  • Takeshi Nomura
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 966)

Abstract

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 words

Bacteria Archaea Cephalexin Lysozyme Mechanosensitive channels K+ channels Patch clamp Laplace’s law 

Notes

Acknowledgments

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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Copyright information

© Springer Science+Business Media New York 2013

Authors and Affiliations

  • Boris Martinac
    • 1
    • 2
    Email author
  • Paul R. Rohde
    • 1
  • Charles G. Cranfield
    • 1
  • Takeshi Nomura
    • 1
  1. 1.Victor Chang Cardiac Research InstituteDarlinghurstAustralia
  2. 2.St. Vincent’s Clinical SchoolUniversity of New South WalesSydneyAustralia

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