Journal
METABOLIC ENGINEERING
Volume 43, Issue -, Pages 21-28Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.ymben.2017.08.001
Keywords
Biosensor; Genetic selection; N-acetylneuraminate; Pathway optimization; NeuAc synthase
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Funding
- National Natural Science Foundation of China [31670077]
- State Key Laboratory of Microbial Technology
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N-acetylneuraminate (NeuAc) biosynthesis has drawn much attention owing to its wide applications in many aspects. Previously, we engineered for the first time an artificial NeuAc biosynthetic pathway in Escherichia coli using glucose as sole substrate. However, rigorous requirements for the flux and cofactor balance make subsequent strain improvement rather difficult. In this study, an in vivo NeuAc biosensor was designed and applied for genetic screening the mutant library of NeuAc producer. Its NeuAc responsive manner was demonstrated using sfgfp as a reporter and a Ni2+-based selection system was developed to couple the cell growth with in vivo NeuAc concentration. Employing this selection system, the NeuAc biosynthesis pathway was optimized and the key enzyme NeuAc synthase was evolved, which improved the titer by 34% and 23%, respectively. The final strain produced up to 8.31 g/L NeuAc in minimal medium using glucose as sole carbon source. This work demonstrated the effectiveness of NeuAc biosensor in genetic screening and great potential in metabolic engineering of other organisms.
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