Combining in vitro translation with nanodisc technology and functional reconstitution of channels in planar lipid bilayers

Abstract

Experimental studies on membrane proteins have been recently enriched by two promising method developments: protocols for cell-free protein synthesis and the use of soluble nanoscale lipid bilayers, so called nanodiscs, as membrane mimics for keeping these proteins in a soluble form. Here, we show how the advantages of these techniques can be combined with the classical planar lipid bilayer method for a functional reconstitution of channel activity. The present data demonstrate that the combination of these methods offers a very rapid and reliable way of recording channel activity in different bilayer systems. This approach has additional advantages in that it strongly lowers the propensity of contamination from the expression system and allows the simultaneous reconstitution of thousands of channel proteins for macroscopic current measurements without compromising bilayer stability.

Keywords: Asymmetric bilayer; Cell-free channel synthesis, nanodisc membrane mimics; Ion channel reconstitution in planar lipid bilayer.