High-yield vesicle-packaged recombinant protein production from E. coli

Tara A. Eastwood; Karen Baker; Bree R. Streather; Nyasha Allen; Lin Wang; Stanley W. Botchway; Ian R. Brown; Jennifer R. Hiscock; Christopher Lennon; Daniel P. Mulvihill

High-yield vesicle-packaged recombinant protein production from E. coli

2023

Summary: We describe an innovative system that exports diverse recombinant proteins in membrane-bound vesicles from E. coli. These recombinant vesicles compartmentalize proteins within a micro-environment that enables production of otherwise challenging insoluble, toxic, or disulfide-bond containing proteins from bacteria. The release of vesicle-packaged proteins supports isolation from the culture and allows long-term storage of active protein. This technology results in high yields of vesicle-packaged, functional proteins for efficient downstream processing for a wide range of applications from discovery science to applied biotechnology and medicine. Motivation: The ability to reprogram a cell to direct the packaging of specific molecules into discrete membrane envelopes is one of the major challenges in the fields of synthetic biology and recombinant protein today. We thus set out to develop a system to allow the export of vesicle-packaged proteins from Escherichia coli. The resultant technology, involving a simple peptide tag, not only simplifies subsequent recombinant protein purification but the controlled packaging into membrane vesicles can be applied to the development of numerous technologies and commercializable products within the biotechnology and medical industries, including generation of recombinant bioreactors, environmental dispersion of biomolecules, and vehicles for drug delivery and vaccination, as well as providing a stable environment for isolation and storage of proteins.

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