Chen, Taichi
(Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, Jiangnan University)
,
Xia, Hongzhi
(Richen Bioengineering Co., Ltd.)
,
Cui, Shixiu
(Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, Jiangnan University)
,
Lv, Xueqin
(Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, Jiangnan University)
,
Li, Xueliang
(Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, Jiangnan University)
,
Liu, Yanfeng
(Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, Jiangnan University)
,
Li, Jianghua
(Key Laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University)
,
Du, Guocheng
(Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, Jiangnan University)
,
Liu, Long
(Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, Jiangnan University)
Vitamin K2 (menaquinone) is an essential vitamin existing in the daily diet, and menaquinone-7 (MK-7) is an important form of it. In a recent work, we engineered the synthesis modules of MK-7 in Bacillus subtilis, and the strain BS20 could produce 360 mg/l MK-7 in shake flasks, while the methyleryth...
Vitamin K2 (menaquinone) is an essential vitamin existing in the daily diet, and menaquinone-7 (MK-7) is an important form of it. In a recent work, we engineered the synthesis modules of MK-7 in Bacillus subtilis, and the strain BS20 could produce 360 mg/l MK-7 in shake flasks, while the methylerythritol phosphate (MEP) pathway, which provides the precursor isopentenyl diphosphate for MK-7 synthesis, was not engineered. In this study, we overexpressed five genes of the MEP pathway in BS20 and finally obtained a strain (BS20DFHG) with MK-7 titer of 415 mg/l in shake flasks. Next, we optimized the fermentation process parameters (initial pH, temperature and aeration) in an 8-unit parallel bioreactor system consisting of 300-ml glass vessels. Based on this, we scaled up the MK-7 production by the strain BS20DFHG in a 50-l bioreactor, and the highest MK-7 titer reached 242 mg/l. Here, we show that the engineered strain BS20DFHG may be used for the industrial production of MK-7 in the future.
Vitamin K2 (menaquinone) is an essential vitamin existing in the daily diet, and menaquinone-7 (MK-7) is an important form of it. In a recent work, we engineered the synthesis modules of MK-7 in Bacillus subtilis, and the strain BS20 could produce 360 mg/l MK-7 in shake flasks, while the methylerythritol phosphate (MEP) pathway, which provides the precursor isopentenyl diphosphate for MK-7 synthesis, was not engineered. In this study, we overexpressed five genes of the MEP pathway in BS20 and finally obtained a strain (BS20DFHG) with MK-7 titer of 415 mg/l in shake flasks. Next, we optimized the fermentation process parameters (initial pH, temperature and aeration) in an 8-unit parallel bioreactor system consisting of 300-ml glass vessels. Based on this, we scaled up the MK-7 production by the strain BS20DFHG in a 50-l bioreactor, and the highest MK-7 titer reached 242 mg/l. Here, we show that the engineered strain BS20DFHG may be used for the industrial production of MK-7 in the future.
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제안 방법
All experiments were independently carried out at least three times, and the results were expressed as mean ± standard deviation (SD).
In this study, we focused on enhancing the transcription level of five isp-genes (ispD, ispE, ispF, ispH, and ispG) in the MEP pathway to promote the production of MK-7. The highest MK-7 production of 415 ± 3.
In a previous work, the dxs, dxr, idi, and menA genes were overexpressed in BS168 to improve the production of MK-7 [5]. In this study, we overexpressed the isp-genes of MEP pathway in the BS20 strain to increase the production of MK-7. We supposed that overexpression of isp-genes in MEP pathway had a positive effect on the supply of IPP and accumulation of MK-7.
성능/효과
In conclusion, after increasing the relative transcription level of isp-genes to increase the flux of the MEP pathway, we obtained the highest production of MK-7 as 415 ± 3.2 mg/l in BS20DFHG, a 29% increase compared with BS20.
Thus, we obtained the highest production of MK-7 as 415 ± 3.2 mg/l (Fig. 2D), a 29% increase compared with BS20 and the relative transcriptional level of ispG in BS20DFHG was 6.9-times that in BS20.
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