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Preparing Protein Producing Synthetic Cells using Cell Free Bacterial Extracts, Liposomes and Emulsion Transfer
Published on: April 27, 2020
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Optimizing Cell-Free Protein Synthesis for Antimicrobial Protein Production
Tejaswini Ramakrishna Hegde1, Ogechi Okocha Rufus2, Joongoo Lee3
1Department of Chemical and Biological Engineering, Illinois Institute of Technology, Chicago, IL, USA.
Methods in Molecular Biology (Clifton, N.J.)
|September 29, 2023
Summary
Cell-free protein synthesis enables production of challenging proteins like colicins, which are antimicrobial proteins with antibiotic potential. This method simplifies the creation of active colicins using molecular chaperones and immunity proteins.
Area of Science:
- Biochemistry
- Molecular Biology
- Microbiology
Background:
- Cell-free protein synthesis (CFPS) offers a versatile method for producing proteins without the need for cell viability.
- Bacteriocins, including colicins, are antimicrobial proteins with significant potential as alternatives to conventional antibiotics.
- Escherichia coli produces colicins that target non-host strains, presenting a model system for studying these agents.
Purpose of the Study:
- To detail protocols for the production and characterization of active colicins using cell-free systems.
- To demonstrate the utility of CFPS for generating difficult-to-express antimicrobial proteins.
- To explore the role of molecular chaperones and immunity proteins in successful colicin production via CFPS.
Main Methods:
- Utilized cell-free protein synthesis reactions.
- Incorporated molecular chaperones into the reaction lysates.
- Co-produced colicin immunity proteins alongside the target colicins.
Main Results:
- Successfully produced active colicins using cell-free extracts.
- Demonstrated that CFPS can yield functional antimicrobial proteins.
- Showcased the importance of chaperones and immunity proteins for efficient colicin synthesis in vitro.
Conclusions:
- Cell-free protein synthesis is an effective platform for producing active colicins.
- This approach facilitates the development of bacteriocins as novel antibiotic alternatives.
- Optimized CFPS protocols can overcome challenges in producing complex antimicrobial proteins.
