ATP and NADPH engineering of Escherichia coli to improve the production of 4-hydroxyphenylacetic acid using CRISPRi

Autor: Yu-Ping Shen, Yu-Ling Liao, Qian Lu, Xin He, Zhi-Bo Yan, Jian-Zhong Liu
Jazyk: angličtina
Rok vydání: 2021
Předmět:
Zdroj: Biotechnology for Biofuels, Vol 14, Iss 1, Pp 1-10 (2021)
Druh dokumentu: article
ISSN: 1754-6834
02475472
DOI: 10.1186/s13068-021-01954-6
Popis: Abstract Background 4-Hydroxyphenylacetic acid (4HPAA) is an important raw material for the synthesis of drugs, pesticides and biochemicals. Microbial biotechnology would be an attractive approach for 4HPAA production, and cofactors play an important role in biosynthesis. Results We developed a novel strategy called cofactor engineering based on clustered regularly interspaced short palindromic repeat interference (CRISPRi) screening (CECRiS) for improving NADPH and/or ATP availability, enhancing the production of 4HPAA. All NADPH-consuming and ATP-consuming enzyme-encoding genes of E. coli were repressed through CRISPRi. After CRISPRi screening, 6 NADPH-consuming and 19 ATP-consuming enzyme-encoding genes were identified. The deletion of the NADPH-consuming enzyme-encoding gene yahK and the ATP-consuming enzyme-encoding gene fecE increased the production of 4HPAA from 6.32 to 7.76 g/L. Automatically downregulating the expression of the pabA gene using the Esa-PesaS quorum-sensing-repressing system further improved the production of 4HPAA. The final strain E. coli 4HPAA-∆yfp produced 28.57 g/L of 4HPAA with a yield of 27.64% (mol/mol) in 2-L bioreactor fed-batch fermentations. The titer and yield are the highest values to date. Conclusion This CECRiS strategy will be useful in engineering microorganisms for the high-level production of bioproducts.
Databáze: Directory of Open Access Journals
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