Affiliation:
1. The National and Local Joint Engineering Research Center for Biomanufacturing of Chiral Chemicals Zhejiang University of Technology Hangzhou Zhejiang P. R. China
2. Key Laboratory of Bioorganic Synthesis of Zhejiang Province College of Biotechnology and Bioengineering Zhejiang University of Technology Hangzhou Zhejiang P. R. China
Abstract
AbstractL‐Cysteine production through fermentation stands as a promising technology. However, excessive accumulation of L‐cysteine poses a challenge due to the potential to inflict damage on cellular DNA. In this study, we employed a synergistic approach encompassing atmospheric and room temperature plasma mutagenesis (ARTP) and adaptive laboratory evolution (ALE) to improve L‐cysteine tolerance in Escherichia coli. ARTP‐treated populations obtained substantial enhancement in L‐cysteine tolerance by ALE. Whole‐genome sequencing, transcription analysis, and reverse engineering, revealed the pivotal role of an effective export mechanism mediated by gene eamB in augmenting L‐cysteine resistance. The isolated tolerant strain, 60AP03/pTrc‐cysEf, achieved a 2.2‐fold increase in L‐cysteine titer by overexpressing the critical gene cysEf during batch fermentation, underscoring its enormous potential for L‐cysteine production. The production evaluations, supplemented with L‐serine, further demonstrated the stability and superiority of tolerant strains in L‐cysteine production. Overall, our work highlighted the substantial impact of the combined ARTP and ALE strategy in increasing the tolerance of E. coli to L‐cysteine, providing valuable insights into improving L‐cysteine overproduction, and further emphasized the potential of biotechnology in industrial production.