A new automated technique for the reconstitution of hydrophobic proteins into planar bilayer membranes. Studies of human recombinant uncoupling protein 1

Biochimica Et Biophysica Acta
Valeri BeckElena E Pohl

Abstract

Electrophysiological characterisation of the vast number of annotated channel and transport proteins in the postgenomic era would be greatly facilitated by the introduction of rapid and robust methods for the functional incorporation of membrane proteins into defined lipid bilayers. Here, we describe an automated technique for reconstitution of membrane proteins into lipid bilayer membranes, which substantially reduces both the reconstitution time and the amount of protein required for the membrane formation. The method allows the investigation of single protein channels as well as insertion of multiple copies (approximately 10(7)) into a single bilayer. Despite a comparatively large membrane area (up to 300 microm diameter), the high stability of the membrane permits the application of transmembrane voltages up to 300 mV. This feature is especially important for studies of inner membrane mitochondrial proteins, since they act at potentials up to approximately 200 mV under physiological conditions. It is a combination of these advantages that enables the detailed investigation of the minuscule single protein conductances typical for proton transporters. We have applied the new technique for the reconstitution and electrophysiol...Continue Reading

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Citations

Aug 26, 2009·Journal of Bioenergetics and Biomembranes·Nadeene ParkerMartin D Brand
Jan 11, 2013·Oxidative Medicine and Cellular Longevity·Petr JežekLydie Plecitá-Hlavatá
Mar 9, 2010·FEBS Letters·Petr JezekKeith D Garlid
Mar 15, 2011·Molecular and Cellular Neurosciences·Alina SmorodchenkoElena E Pohl
Nov 1, 2015·Biochimica Et Biophysica Acta·Karolina E HilseElena E Pohl
Mar 26, 2016·Free Radical Biology & Medicine·Andrea AnnibalMaria Fedorova
Aug 19, 2007·Biochimica Et Biophysica Acta·Inna I SeverinaVladimir P Skulachev
Feb 11, 2015·Analytical and Bioanalytical Chemistry·Sandra MarxenThomas Hofmann
Mar 4, 2008·Biochimica Et Biophysica Acta·Elena E PohlAnne Rupprecht
Apr 23, 2010·Biophysical Journal·Anne RupprechtElena E Pohl
Jan 21, 2018·Antioxidants & Redox Signaling·Petr JežekMartin Jabůrek
Jan 24, 2007·FASEB Journal : Official Publication of the Federation of American Societies for Experimental Biology·Valeri BeckElena E Pohl
Feb 4, 2021·International Journal of Molecular Sciences·Sanja ŠkuljMario Vazdar
Apr 4, 2021·International Journal of Molecular Sciences·Jürgen KreiterElena E Pohl

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