Ye Runle, Li Feng, Ding Fan, Zhao Zhenhui, Chen Sheng, Yuan Jianfeng
Xingzhi College, Zhejiang Normal University, Lanxi 321100, Zhejiang, China.
Key Laboratory of Wildlife Biotechnology and Conservation and Utilization of Zhejiang Province, Zhejiang Normal University, Jinhua 321004, Zhejiang, China.
Sheng Wu Gong Cheng Xue Bao. 2020 Jun 25;36(6):1138-1149. doi: 10.13345/j.cjb.190482.
Pyrroloquinoline quinone (PQQ), an important redox enzyme cofactor, has many physiological and biochemical functions, and is widely used in food, medicine, health and agriculture industry. In this study, PQQ production by recombinant Gluconobacter oxydans was investigated. First, to reduce the by-product of acetic acid, the recombinant strain G. oxydans T1 was constructed, in which the pyruvate decarboxylase (GOX1081) was knocked out. Then the pqqABCDE gene cluster and tldD gene were fused under the control of endogenous constitutive promoter P0169, to generate the recombinant strain G. oxydans T2. Finally, the medium composition and fermentation conditions were optimized. The biomass of G. oxydans T1 and G. oxydans T2 were increased by 43.02% and 38.76% respectively, and the PQQ production was 4.82 and 20.5 times higher than that of the wild strain, respectively. Furthermore, the carbon sources and culture conditions of G. oxydans T2 were optimized, resulting in a final PQQ yield of (51.32±0.899 7 mg/L), 345.6 times higher than that of the wild strain. In all, the biomass of G. oxydans and the yield of PQQ can be effectively increased by genetic engineering.
吡咯喹啉醌(PQQ)是一种重要的氧化还原酶辅因子,具有多种生理生化功能,广泛应用于食品、医药、保健和农业等行业。本研究对重组氧化葡萄糖酸杆菌生产PQQ进行了探究。首先,为减少副产物乙酸的生成,构建了敲除丙酮酸脱羧酶(GOX1081)的重组菌株氧化葡萄糖酸杆菌T1。然后,将pqqABCDE基因簇和tldD基因在内源组成型启动子P0169的控制下融合,构建出重组菌株氧化葡萄糖酸杆菌T2。最后,对培养基成分和发酵条件进行了优化。氧化葡萄糖酸杆菌T1和氧化葡萄糖酸杆菌T2的生物量分别提高了43.02%和38.76%,PQQ产量分别是野生菌株的4.82倍和20.5倍。此外,对氧化葡萄糖酸杆菌T2的碳源和培养条件进行了优化,最终PQQ产量达到(51.32±0.899 7 mg/L),是野生菌株的345.6倍。总之,通过基因工程可有效提高氧化葡萄糖酸杆菌的生物量和PQQ产量。