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  Introduction to Solid Supported Membrane Based Electrophysiology

Bazzone, A., Costa, W. S., Braner, M., Călinescu, O., Hatahet, L., & Fendler, K. (2013). Introduction to Solid Supported Membrane Based Electrophysiology. Journal of Visualized Experiments, 75: e50230. doi:10.3791/50230.

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 Creators:
Bazzone, Andre1, Author           
Costa, Wagner Steuer2, Author
Braner, Markus1, Author           
Călinescu, Octavian1, Author           
Hatahet, Lina1, Author           
Fendler, Klaus1, Author           
Affiliations:
1Department of Biophysical Chemistry, Max Planck Institute of Biophysics, Max Planck Society, ou_2068289              
2Buchmann Institute for Molecular Life Sciences, Goethe-University Frankfurt, ou_persistent22              

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Free keywords: Biochemistry; Issue 75; Biophysics; Molecular Biology; Cellular Biology; Physiology; Proteins; Membrane Lipids; Membrane Transport; Proteins; Kinetics; Electrophysiology; solid supported membrane; SSM; membrane transporter; lactose permease; lacy; capacitive coupling; solution exchange; model membrane; membrane protein; transporter; kinetics; transport mechanism
 Abstract: The electrophysiological method we present is based on a solid supported membrane (SSM) composed of an octadecanethiol layer chemisorbed on a gold coated sensor chip and a phosphatidylcholine monolayer on top. This assembly is mounted into a cuvette system containing the reference electrode, a chlorinated silver wire. After adsorption of membrane fragments or proteoliposomes containing the membrane protein of interest, a fast solution exchange is used to induce the transport activity of the membrane protein. In the single solution exchange protocol two solutions, one non-activating and one activating solution, are needed. The flow is controlled by pressurized air and a valve and tubing system within a faraday cage. The kinetics of the electrogenic transport activity is obtained via capacitive coupling between the SSM and the proteoliposomes or membrane fragments. The method, therefore, yields only transient currents. The peak current represents the stationary transport activity. The time dependent transporter currents can be reconstructed by circuit analysis. This method is especially suited for prokaryotic transporters or eukaryotic transporters from intracellular membranes, which cannot be investigated by patch clamp or voltage clamp methods.

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Language(s): eng - English
 Dates: 2013-05-11
 Publication Status: Published online
 Pages: 11
 Publishing info: -
 Table of Contents: -
 Rev. Type: Peer
 Identifiers: DOI: 10.3791/50230
PMID: 23711952
PMC: PMC3679796
 Degree: -

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Title: Journal of Visualized Experiments
  Other : Journal of Visualized Experiments: JoVE
  Abbreviation : J. Vis. Exp.
Source Genre: Journal
 Creator(s):
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Publ. Info: Rockville Pike, Bethesda MD : JoVE
Pages: - Volume / Issue: 75 Sequence Number: e50230 Start / End Page: - Identifier: ISSN: 1940-087X
CoNE: https://pure.mpg.de/cone/journals/resource/1940087X