Engineering Escherichia coli for l-homoserine production.

Autor: Sun BY; State Key Lab of Bioreactor Engineering, Newworld Institute of Biotechnology, East China University of Science and Technology, Shanghai, China., Wang FQ; State Key Lab of Bioreactor Engineering, Newworld Institute of Biotechnology, East China University of Science and Technology, Shanghai, China., Zhao J; State Key Lab of Bioreactor Engineering, Newworld Institute of Biotechnology, East China University of Science and Technology, Shanghai, China., Tao XY; State Key Lab of Bioreactor Engineering, Newworld Institute of Biotechnology, East China University of Science and Technology, Shanghai, China., Liu M; State Key Lab of Bioreactor Engineering, Newworld Institute of Biotechnology, East China University of Science and Technology, Shanghai, China., Wei DZ; State Key Lab of Bioreactor Engineering, Newworld Institute of Biotechnology, East China University of Science and Technology, Shanghai, China.
Jazyk: angličtina
Zdroj: Journal of basic microbiology [J Basic Microbiol] 2023 Feb; Vol. 63 (2), pp. 168-178. Date of Electronic Publication: 2022 Oct 25.
DOI: 10.1002/jobm.202200488
Abstrakt: l-homoserine, a nonprotein amino acid, is used to synthesize many active substances in the industry. Here, to develop a robust l-homoserine-producing strain, Escherichia coli W3110 was used as a chassis to be engineered. Based on a previous construct with blocked competing routes for l-homoserine synthesis, five genes were overexpressed by promoter replacement strategy to increase the l-homoserine production, including enhancement of precursors for l-homoserine synthesis (ppc, thrA, and asd), reinforcement of the NADPH supply (pntAB) and efflux transporters (rhtA) to improve the l-homoserine production. However, the plasmid losing was to blame for the wildly fluctuating fermentation performance of engineered strains, ranging between 2.1 and 6.2 g/L. Then, a hok/sok toxin/antitoxin system was introduced into the free plasmid expression cassette to maintain the genetic stability of the episomal plasmid; consequently, the plasmid-losing rate sharply decreased, resulting in the engineered strain SHL17, which exhibited excellent stability in l-homoserine production, with 6.3 g/L in shake flasks and 44.4 g/L in a 5-L fermenter without antibiotic addition. This work verified the effective use of the hok/sok toxin/antitoxin system combined with promoter engineering to improve the genetic stability of E. coli episomal plasmids without antibiotics.
(© 2022 Wiley-VCH GmbH.)
Databáze: MEDLINE
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