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A well-characterized polycistronic-like gene expression system in yeast
Minakshi Mukherjee1, Zhen Q Wang1
1Department of Biological Sciences, University at Buffalo, The State University of New York, Buffalo, New York, USA.
Biotechnology and Bioengineering
|September 28, 2022
Summary
This study introduces a novel yeast polycistronic expression system using 2A viral peptides and modular cloning for efficient multigene expression. The system successfully produced geraniol, demonstrating its utility in metabolic engineering for chemical bioproduction.
Area of Science:
- Synthetic Biology
- Metabolic Engineering
- Molecular Biology
Background:
- Efficient multigene expression is crucial for yeast metabolic engineering.
- Polycistronic expression systems, driven by a single promoter, offer advantages for bioproduction.
- 2A viral peptides facilitate co-translational expression of multiple proteins via ribosomal skipping.
Purpose of the Study:
- To develop and characterize a modular cloning (MoClo) compatible 2A peptide-based polycistronic-like system for yeast.
- To assess the efficiency of different 2A peptides (E2A, P2A, O2A) in yeast.
- To evaluate the system's performance in producing a valuable industrial compound, geraniol.
Main Methods:
- Developed a MoClo-compatible system for assembling polycistronic constructs.
- Co-expressed fluorescent proteins in bi-, tri-, and quad-cistronic configurations to assess 2A peptide cleavage efficiency.
- Engineered yeast strains using the polycistronic system for geraniol production.
Main Results:
- Demonstrated high cleavage efficiencies for E2A, P2A, and O2A peptides in yeast.
- Achieved comparable or higher geraniol titers using the polycistronic system versus monocistronic constructs.
- Validated the system's effectiveness for multigene expression in metabolic engineering.
Conclusions:
- The developed 2A peptide-based polycistronic-like system is a valuable tool for yeast metabolic engineering.
- This system facilitates efficient multigene expression from a single promoter.
- It offers a streamlined approach for the bioproduction of chemicals.
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