School of Life Sciences, Institute of Aquatic Economic Animals and Guangdong Provincial Key Laboratory for Aquatic Economic Animals, National Engineering Center for Marine Biotechnology of South China Sea, Sun Yat-Sen University , Guangzhou, Guangdong 510275, P. R. China.
Dongguan Agricultural Research Center , Dongguan, Guangdong 523086, P. R. China.
J Agric Food Chem. 2017 Dec 13;65(49):10757-10766. doi: 10.1021/acs.jafc.7b04694. Epub 2017 Dec 5.
In this study, three kinds of milk were treated with the β-galactosidase Bgal1-3 (4 U/mL), resulting in 7.2-9.5 g/L galactooligosaccharides (GOS) at a lactose conversion of 90-95%. Then, Bgal1-3 was secreted from Pichia pastoris X33 under the direction of an α-factor signal peptide. After cultivation for 144 h in a flask culture with shaking, the extracellular activity of Bgal1-3 was 4.4 U/mL. Five more signal peptides (HFBI, apre, INU1A, MF4I, and W1) were employed to direct the secretion, giving rise to a more efficient signal peptide, W1 (11.2 U/mL). To further improve the secretion yield, recombinant strains harboring two copies of the bgal1-3 gene were constructed, improving the extracellular activity to 22.6 U/mL (about 440 mg/L). This study successfully constructed an engineered strain for the production of the β-galactosidase Bgal1-3, which is a promising catalyst in the preparation of prebiotic-enriched milk.
在这项研究中，三种牛奶经β-半乳糖苷酶 Bgal1-3（4 U/mL）处理，乳糖转化率为 90-95%时可得到 7.2-9.5 g/L 的半乳糖低聚糖（GOS）。然后，毕赤酵母 X33 在α-因子信号肽的指导下分泌 Bgal1-3。在摇瓶培养中培养 144 h 后，Bgal1-3 的细胞外活性为 4.4 U/mL。另外 5 种信号肽（HFBI、apre、INU1A、MF4I 和 W1）被用来指导分泌，其中 W1（11.2 U/mL）是一种更有效的信号肽。为了进一步提高分泌产量，构建了携带两个 bgal1-3 基因拷贝的重组菌株，将细胞外活性提高到 22.6 U/mL（约 440 mg/L）。本研究成功构建了用于生产β-半乳糖苷酶 Bgal1-3 的工程菌株，该酶是制备富含益生元牛奶的有前途的催化剂。
