National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, 1800 Lihu Road, Wuxi, Jiangsu 214122, China; School of Biotechnology and Key Laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University, 1800 Lihu Road, Wuxi, Jiangsu 214122, China; Science Center for Future Foods, Jiannan University, 1800 Lihu Road, Wuxi, Jiangsu 214122, China.
National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, 1800 Lihu Road, Wuxi, Jiangsu 214122, China; School of Biotechnology and Key Laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University, 1800 Lihu Road, Wuxi, Jiangsu 214122, China; Science Center for Future Foods, Jiannan University, 1800 Lihu Road, Wuxi, Jiangsu 214122, China.
Enzyme Microb Technol. 2021 Oct;150:109863. doi: 10.1016/j.enzmictec.2021.109863. Epub 2021 Jun 29.
Protein-glutaminase (EC 3.5.1.44, PG) converts protein glutamine residues in proteins and peptides into glutamic acid residue, and markedly improves the solubility, emulsification, and foaming properties of food proteins. However, the source bacteria, Chryseobacterium proteolyticum, have low enzyme production ability, inefficient genetic operation, and high production cost. Therefore, it is critical to establish an efficient expression system for active PG. Here, combinatorial engineering was developed for high-yield production of PG in Bacillus subtilis. First, we evaluated different B. subtilis strains for PG self-activation. Then, combinatorial optimization involving promoters, signal peptides, and culture medium was applied to produce active recombinant PG in a B. subtilis expression system. Through combinatorial engineering, PG enzyme activity reached 3.23 U/mL in shaken-flask cultures. Active PG with the yield of 7.07 U/mL was obtained at 40 h by the P-YdeJ combination in fed-batch fermentation, which is the highest yield of PG in existing reports.
蛋白谷氨酰胺酶（EC 3.5.1.44，PG）可将蛋白质和肽中的谷氨酰胺残基转化为谷氨酸残基，显著提高食品蛋白的溶解度、乳化性和起泡性。然而，产酶菌希瓦氏菌（Chryseobacterium proteolyticum）的产酶能力低、遗传操作效率低、生产成本高。因此，建立高效的 PG 活性表达系统至关重要。本研究采用组合工程技术在枯草芽孢杆菌中高效生产 PG。首先，我们评估了不同枯草芽孢杆菌菌株对 PG 自我激活的能力。然后，通过组合优化启动子、信号肽和培养基，在枯草芽孢杆菌表达系统中生产具有活性的重组 PG。通过组合工程，摇瓶培养中 PG 酶活达到 3.23 U/mL。在补料分批发酵中，采用 P-YdeJ 组合，在 40 h 时获得了 7.07 U/mL 的活性 PG，这是现有报道中 PG 的最高产量。
