Abstract
Hybrid membranes comprised of diblock copolymers, and phospholipids have gained interest due to their unique properties that result from blending natural and synthetic components. The integration of membrane proteins into these synthetic membranes is an important step towards creating biomembrane systems for uses such as artificial cellular systems, biosensors, and drug delivery vehicles. Here, we outline a technique to create hybrid membranes composed of phospholipids and diblock copolymers. Next, we describe how membrane proteins can be co-translationally integrated into hybrid lipid/polymer membranes using a cell-free reaction. We then outline a method to monitor insertion and folding of a membrane-embedded channel protein into the hybrid membrane using a fluorescent-protein reporter and dye release assay, respectively. This method is expected to be applicable for a wide range of membrane proteins that do not require chaperones for co-translational integration into vesicles and provides a generalized protocol for expressing a membrane protein into a membrane mimetic.
Key words
- Membrane protein
- Vesicles
- Cell-free protein synthesis
- Diblock copolymer
- Hybrid membranes
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Acknowledgments
This work was supported by the Air Force Office of Scientific Research (AFOSR) YIP FA9550-19-1-0039 P00001 to NPK and NSF grant MCB-1935356 (NPK). M.L.J. was supported by Grant No. T32GM008382 from the National Institute of General Medical Sciences and The American Heart Association Predoctoral Fellowship under Grant No. 20PRE35180215.
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Jacobs, M.L., Kamat, N.P. (2022). Cell-Free Membrane Protein Expression into Hybrid Lipid/Polymer Vesicles. In: Karim, A.S., Jewett, M.C. (eds) Cell-Free Gene Expression. Methods in Molecular Biology, vol 2433. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1998-8_16
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DOI: https://doi.org/10.1007/978-1-0716-1998-8_16
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Online ISBN: 978-1-0716-1998-8
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