When recombinant E.coli BL21(DE3)is induced to express Bacillus subtilis lipase A(BsLipA)using Isopropylβ-D-1-thiogalactopyranoside(IPTG),the one-time addition of IPTG leads to problems such as limited cell growth,a ...When recombinant E.coli BL21(DE3)is induced to express Bacillus subtilis lipase A(BsLipA)using Isopropylβ-D-1-thiogalactopyranoside(IPTG),the one-time addition of IPTG leads to problems such as limited cell growth,a short enzyme production period,and low yield.To address these issues,this study proposes an innovative IPTG feeding strategy,where the IPTG feeding rate is adjusted based on cell growth rate between 4 and 10 h,followed by a constant IPTG feeding rate after 12 h.Fermentation experiments in a 5 L bioreactor demonstrated that IPTG feeding according to this strategy resulted in continuous enhancement of BsLipA activity,reaching 288.50 U/mL.Compared to a batch induced with a one-time addition of 0.2 mmol/L IPTG,BsLipA activity increased by 6.67 times.This IPTG feeding strategy was applied to a low-nutrient fermentation process with DO-start glucose feeding,leading to a further increase in BsLipA enzyme activity,with the highest activity reaching 580.29 U/mL.The results indicate that this strategy significantly reduces the toxic effects of IPTG on the cells,improves biomass,extends the enzyme production phase,and enhances BsLipA expression levels by balancing the induction strength and cell growth conditions.展开更多
基金funded by the Jiangsu Basic Research Center for Synthetic Biology(Grant No.BK20233003)the Key Project of the Bayannur National Agricultural High-Tech Industry Demonstration Zone(Grant Nos.NMKJXM202210,NMKJXM202309).
文摘When recombinant E.coli BL21(DE3)is induced to express Bacillus subtilis lipase A(BsLipA)using Isopropylβ-D-1-thiogalactopyranoside(IPTG),the one-time addition of IPTG leads to problems such as limited cell growth,a short enzyme production period,and low yield.To address these issues,this study proposes an innovative IPTG feeding strategy,where the IPTG feeding rate is adjusted based on cell growth rate between 4 and 10 h,followed by a constant IPTG feeding rate after 12 h.Fermentation experiments in a 5 L bioreactor demonstrated that IPTG feeding according to this strategy resulted in continuous enhancement of BsLipA activity,reaching 288.50 U/mL.Compared to a batch induced with a one-time addition of 0.2 mmol/L IPTG,BsLipA activity increased by 6.67 times.This IPTG feeding strategy was applied to a low-nutrient fermentation process with DO-start glucose feeding,leading to a further increase in BsLipA enzyme activity,with the highest activity reaching 580.29 U/mL.The results indicate that this strategy significantly reduces the toxic effects of IPTG on the cells,improves biomass,extends the enzyme production phase,and enhances BsLipA expression levels by balancing the induction strength and cell growth conditions.
