Compartments for the spatially and temporally controlled assembly of biological processes are essential towards cellular life. Synthetic mimics of cellular compartments based on lipid-based protocells lack the mechanical and chemical stability to allow their manipulation into a complex and fully functional synthetic cell. Here, we present a high-throughput microfluidic method to generate stable, defined sized liposomes termed ‘droplet-stabilized giant unilamellar vesicles (dsGUVs)’. The enhanced stability of dsGUVs enables the sequential loading of these compartments with biomolecules, namely purified transmembrane and cytoskeleton proteins by microfluidic pico-injection technology. This constitutes an experimental demonstration of a successful bottom-up assembly of a compartment with contents that would not self-assemble to full functionality when simply mixed together. Following assembly, the stabilizing oil phase and droplet shells are removed to release functional self-supporting protocells to an aqueous phase, enabling them to interact with physiologically relevant matrices.
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Parts of the research leading to these results have received funding from the European Research Council/ERC Grant Agreement no. 294852, SynAd. This work is also part of the MaxSynBio consortium, which is jointly funded by the Federal Ministry of Education and Research of Germany and the Max Planck Society. The work was also partly supported by the SFB 1129 of the German Science Foundation and the VolkswagenStiftung (priority call ‘Life?’). J.P.S. is the Weston Visiting Professor at the Weizmann Institute of Science and part of the excellence cluster CellNetworks at the University of Heidelberg. J.-C.B. acknowledges financial support by the ERC (FP7/2007-2013/ERC Grant agreement 306385-SofI). I.P. acknowledges the support of the Alexander von Humboldt Foundation. The authors acknowledge the help of P. Gruner and B. Riechers for their technical assistance with preliminary microfluidic experiments and A. Richter (WITec GmbH, Germany) for her technical assistance with Raman microscopy. The support of N. Grunze for editing the manuscript as well as of J. Ricken and Ch. Mollenhauer for their general support in protein purification and chemical synthesis is highly acknowledged. The Max Planck Society is appreciated for its general support in all aspects of our research.
The authors declare no competing financial interests.
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Weiss, M., Frohnmayer, J., Benk, L. et al. Sequential bottom-up assembly of mechanically stabilized synthetic cells by microfluidics. Nature Mater 17, 89–96 (2018). https://doi.org/10.1038/nmat5005
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