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Rapid and Continuous Directed Evolution in Vibrio natriegens Utilizing an In Vivo Hypermutation System
Ruiying Zhu1,2,3, Zehua Bao1,2,3,4
1Key Laboratory of Biomass Chemical Engineering of Ministry of Education, College of Chemical and Biological Engineering, Zhejiang University, Hangzhou, Zhejiang 310058, China.
ACS Synthetic Biology
|June 26, 2026
Summary
We established MutaT7 in Vibrio natriegens, the fastest-growing bacterium, for rapid protein engineering. This accelerated platform achieved high-level ceftazidime resistance in the TEM-1 protein within 36 hours.
Area of Science:
- Synthetic biology
- Molecular evolution
- Microbial biotechnology
Background:
- Genetic diversification is key for in vivo molecular evolution.
- MutaT7 facilitates efficient DNA diversification in E. coli and S. cerevisiae.
- Vibrio natriegens is a fast-growing bacterium, ideal for synthetic biology.
Purpose of the Study:
- Establish MutaT7 in V. natriegens for accelerated protein engineering.
- Demonstrate rapid evolution of TEM-1 protein for ceftazidime resistance.
Main Methods:
- Fusion of cytidine deaminase with T7 RNA polymerase (MutaT7).
- Introduction and establishment of MutaT7 in V. natriegens (VnMutaT7).
- Directed evolution of TEM-1 protein for antibiotic resistance.
Main Results:
- Successful establishment of VnMutaT7 in V. natriegens.
- Achieved high-level ceftazidime resistance in TEM-1 protein.
- Rapid evolution completed in just 36 hours.
Conclusions:
- Vibrio natriegens is a potent chassis for accelerated protein engineering.
- VnMutaT7 enables rapid molecular evolution in fast-growing bacteria.
- This platform has significant potential for synthetic biology applications.
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