代谢工程改造枯草芽孢杆菌促进L-赖氨酸高效合成研究
Metabolic engineering for improving the L-lysine production byBacillus subtilis
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摘要:为了构建具有益生功能和L-赖氨酸合成功能的“双功能”枯草芽孢杆菌( Bacillus subtilis)重组菌株,对饲料工业常用的益生菌 B.subtilisACCC11025进行系统的代谢工程改造。结果表明:以来源于谷氨酸棒杆菌( Corynebacterium glutamicum)的 lysC311、zwf 234和 gnd 361替换 B.subtilis中的 thrD、zwf和gnd,即构建重组菌 B.subtilisXH4,有利于L-赖氨酸的合成,其产量达到(20.3±1.9) g/L;将 B.subtilis中 hom替换成来源于 C.glutamicum的 hom 59,即构建重组菌 B.subtilisXH5,可显著降低副产物积累量,提高L-赖氨酸产量至(23.2±1.7) g/L,且不影响菌体生长;在重组菌 B.subtilisXH5中引入 C.glutamicum中的DapDH会改变二氨基庚二酸途径(DAP)碳分布进而促进L-赖氨酸的合成,目标重组菌 B.subtilisXH6的L-赖氨酸产量达到(25.6±2.3) g/L。
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关键词:
- 枯草芽孢杆菌/
- L-赖氨酸合成/
- CRISPR-Cas9/
- 反馈调节/
- 代谢工程
Abstract:In order to construct a dual-functional B.subtiliswith probiotic function and L-lysine production, B.subtilisACCC11025 was systematically modified. These results indicated that the strain with replacement of thrD,zwfand gndfrom B.subtilisby lysC311,zwf 234and gnd 361from C.glutamicum(i.e., B.subtilisXH4) was beneficial to L-lysine production, and the yield of L-lysine was (20.3±1.9) g/L. In addition, the strain with replacement of homfrom B.subtilisby hom 59from C.glutamicum(i.e., B.subtilisXH5)produced (23.2±1.7) g/L of L-lysine without the decrease of cell growth. In addition, the yield of by-products in B.subtilisXH5 was significantly decreased. Moreover, the DapDH from C.glutamicumwas introduced into the B.subtilisXH5 (i.e, B.subtilisXH6), resulting in the increase of L-lysine production because of the redirection of the carbon flux in DAP pathway. The resulted recombinant strain B.subtilisXH6 produced (25.6±2.3) g/L of L-lysine. -
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