Improving l-serine formation by Escherichia coli by reduced uptake of produced l-serine

Abstract

Background: Microbial de novo production of l-serine, which is widely used in a range of cosmetic and pharmaceutical products, has attracted increasing attention due to its environmentally friendly characteristics. Previous pioneering work mainly focused on l-serine anabolism; however, in this study, it was found that l-serine could be reimported through the l-serine uptake system, thus hampering l-serine production. Result: To address this challenge, engineering via deletion of four genes, namely, sdaC, cycA, sstT and tdcC, which have been reported to be involved in l-serine uptake in Escherichia coli, was first carried out in the l-serine producer E. coli ES. Additionally, the effects of these genes on l-serine uptake activity and l-serine production were investigated. The data revealed an abnormal phenomenon regarding serine uptake activity. The serine uptake activity of the ΔsdaC mutant was 0.798 nmol min −1 (mg dry weight) −1 after 30 min, decreasing by 23.34% compared to that of the control strain. However, the serine uptake activity of the single sstT, cycA and tdcC mutants increased by 34.29%, 78.29% and 48.03%, respectively, compared to that of the control strain. This finding may be the result of the increased level of sdaC expression in these mutants. In addition, multigene-deletion strains were constructed based on an sdaC knockout mutant. The ΔsdaCΔsstTΔtdcC mutant strain exhibited 0.253 nmol min −1 (mg dry weight) −1 l-serine uptake activity and the highest production titer of 445 mg/L in shake flask fermentation, which was more than threefold the 129 mg/L production observed for the parent. Furthermore, the ΔsdaCΔsstTΔtdcC mutant accumulated 34.8 g/L l-serine with a yield of 32% from glucose in a 5-L fermenter after 36 h. Conclusion: The results indicated that reuptake of l-serine impairs its production and that an engineered cell with reduced uptake can address this problem and improve the production of l-serine in E. coli.

Fitness of Chassis Cells and Metabolic Pathways for l-Cysteine Overproduction in Escherichia coli

“…However, some Cys degradation genes are also involved in the synthesis of methionine ( metC and malY ) and Cys ( cysK and cysM ). In addition, because the main Ser deaminase among the four Ser degradation genes ( sdaA , sdaB , tdcG , tdcB ) was identified, , sdaA was often deleted alone to improve Ser formation. − To accumulate more Cys, the effects of deletion of yhaM , tnaA , and sdaA in LH2A1M were investigated. Therefore, LH2A1MΔY (Δ yhaM )-pLH03, LH2A1MΔT (Δ tnaA )-pLH03, and LH2A1MΔS (Δ sdaA )-pLH03 were constructed.…”

Fitness of chassis cells and metabolic pathways for L-cysteine overproduction in Escherichia coli

“…However, some Cys degradation genes are also involved in the synthesis of methionine (metC and malY ) and Cys (cysK and cysM ). In addition, since the main Ser deaminase among the four Ser degradation genes (sdaA , sdaB ,tdcG , tdcB ) was identified so far (Su et al, 1989;Zhang et al, 2008), SdaA was often deleted alone to improve Ser formation (Wang et al, 2020;Y. Zhang et al, 2018;Gu et al, 2014).…”

Transcriptome Analysis Reveals Potential Mechanisms of L-Serine Production by Escherichia coli Fermentation in Different Carbon–Nitrogen Ratio Medium

“…We referred to the method of Wang et al [ 10 ] to determine the bacterial concentration. After turning on the UV-Vis spectrophotometer (DU730, Beckman, Krefeld, Germany), it needed to warm up for 20 min, and then the wavelength for analysis was selected.…”

“…The activated bacterial broth was inoculated into conical flasks containing 50 mL of shake flask fermentation medium at an 8% transfer volume for shake flask fermentation, incubation temperature 33 °C and speed 200 rpm. After 3 h of incubation, the temperature was increased to 38 °C to induce the production of L-serine by the bacteria [ 10 ].…”