Metabolic engineering of Escherichia coli for the synthesis of menaquinone-7
Menaquinone-7(MK-7),as one of the vitamin K2 subtypes,has the effects of treating osteoporosis and protecting cardio-vascular health.Common MK-7 production is mainly fermented by food such as natto,but there are problems such as low yield and com-plex extraction.Bacillus subtilis,as a common production strain of MK-7,also faces the problems of long fermentation period and low titer.In this study,a strain of Escherichia coli introducing mevalonate pathway was modified.Firstly,four key enzymes were screened by DLKcat model,including heptaisopentadiene pyrophosphate synthase BtHepS \ BtHepT,1,4-dihydroxy-2-naphthoic acid heptaenyl transferase MnMenA,and menadione biosynthesis C-methyltransferase NtMenG.The expression optimization was carried out to construct the endoge-nous synthesis of MK-7 pathway in E.coli.Subsequently,the prenylated pyrophosphate isomerase EcIdi was integrated and expressed to enhance the precursor supply,and then the fed-batch fermentation conditions of the obtained strains were optimized,including delaying the induction time,increasing the dissolved oxygen to 60.The MK-7 production reached 929.58 mg/L,but there was 1 217.58 mg/L demethylmenaquinone-7(DMK-7)accumulation.Then,different sources of MenG were further screened to reduce the accumulation of DMK-7,and it was found that MenG from B.subtilis 168 could reduce the accumulation of DMK-7.Finally,the production of MK-7 reached 1 090.45 mg/L and DMK-7 decreased to 231.14 mg/L in a 5 L fermenter.The results of this study have certain guiding signifi-cance for the application of E.coli to produce MK-7.
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