SecYEG-mediated Translocation in a Model Synthetic Cell

Giant unilamellar vesicles (GUVs) provide a powerful model compartment for synthetic cells. However, a key challenge is the incorporation of membrane proteins that allow for transport, energy transduction, compartment growth and division. Here, we have successfully incorporated the membrane protein complex SecYEG – the key bacterial translocase that is essential for the incorporation of newly synthesized membrane proteins – in GUVs. Our method consists of fusion of small unilamellar vesicles (SUVs) containing reconstituted SecYEG into GUVs, thereby forming SecGUVs. These are suitable for large scale experiments while maintaining a high protein:lipid ratio. We demonstrate that incorporation of SecYEG into GUVs does not inhibit its translocation efficiency. Robust membrane protein functionalized proteo-GUVs are promising and flexible compartments for use in the formation and growth of synthetic cells.