Cell-free expression & nanodiscs: The perfect match for functional membrane proteins

Expressing and purifying membrane proteins is still a difficult task, even if only small quantities are required e.g. for interaction studies. Our cell-free lysates can be combined with nanodiscs for an elegant way to produce membrane proteins. Since the effect of phospholipids on membrane protein activity can be significant, we recommend to test a number of different lipids and combinations. With cell-free setups, screening of up to 48 reactions can now be done in parallel in 24 hours, using standard laboratory equipment.

This method has been developed at the University of Frankfurt and is backed up by a number of publications.

 

  Nanodiscs and cell-free expression: the perfect match for functional membrane proteins

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Functional membrane proteins obtained from E.coli cell-free lysates

 

Differential ligand binding activity of the human endothelin A (ETA) and endothelin B (ETB) receptors in MSP1E3D1 (DMPC) nanodiscs

 
 
 
 
 
Fig. 1. Differential ligand binding activity of the human endothelin A (ETA) and endothelin B (ETB) receptors in MSP1E3D1 (DMPC) nanodiscs. Two GPCRs,  ETA and ETB, were synthesized in cell-free lysates supplemented with pre-assembled MSP1E3D1-DMPC nanodiscs. Ligand binding of the resulting receptor/nanodisc to four different peptide ligands was analyzed by SPR (Proverbio et al., (2013) BBA 1828, 2182-2192).  Ligand binding profiles were in good agreement with data from the literature. Data kindly provided by Frank Bernhard, University Frankfurt.
 
Literature reference (in German:)
Frank Bernhard, Erik Henrich, Barbara Maertens: Funktionale Membranproteine durch optimierte Lipidumgebung in Nanodiscs, October 2015, Volume 21, Issue 6, pp 640-642