Key Laboratory of China National Light Industry and Food Bioengineering, College of Food Science and Nutritional Engineering, China Agricultural University, No. 17 Qinghua East Road, Haidian District, Beijing 100083, China.
Mengniu Hi-tech Dairy Product Beijing Co., Ltd., Beijing 101100, China.
J Agric Food Chem. 2024 Nov 27;72(47):26431-26440. doi: 10.1021/acs.jafc.4c07162. Epub 2024 Nov 14.
galacto-oligosaccharide (GOS) synthesis in milk using β-galactosidases is an effective method for developing prebiotic dairy products. However, the low lactose concentration in milk (∼4.6%, w/w) reduces the GOS yield. In this study, a modified β-galactosidase from (mBgaD-D) with enhanced transglycosylation activity at low lactose concentration was developed through directed evolution and saturation mutagenesis. The GOS yield by mBgaD-D increased from 22.8% (wild type) to 30.8% in 50 g/L lactose (phosphate buffer). P was a strong sorbitol-inducible promoter from . The expression of mBgaD-D in , coupled with the P promoter, resulted in a 6.4-fold increase (compared to the P promoter) in extracellular enzyme activity. Additionally, adding whey powder to boost the initial lactose concentration further improved the GOS yield, which reached 43% under the optimized conditions. Combining mBgaD-D and whey powder enhanced milk sweetness, producing no sugar-added, GOS-enriched yogurt (GOSY). The GOS content in GOSY was 4.1/100 g, providing an appropriate level of sweetness and yielding a yogurt that is elastic as well as firm. GOSY also increased the population of spp. during a 24 h fecal fermentation. Thus, fortifying yogurt with mBgaD-D and whey powder can enhance its technological properties and health benefits.
利用β-半乳糖苷酶在牛奶中合成半乳糖寡糖（GOS）是开发益生元乳制品的有效方法。然而，牛奶中乳糖浓度较低（约 4.6%，w/w）会降低 GOS 的产量。本研究通过定向进化和饱和诱变，开发了一种具有低乳糖浓度下增强的转糖苷活性的改良β-半乳糖苷酶（mBgaD-D）。在 50 g/L 乳糖（磷酸盐缓冲液）中，mBgaD-D 的 GOS 产量从 22.8%（野生型）提高到 30.8%。P 是来自 的强山梨醇诱导启动子。mBgaD-D 在 中的表达与 P 启动子结合，使细胞外酶活性提高了 6.4 倍（与 P 启动子相比）。此外，添加乳清粉以提高初始乳糖浓度进一步提高了 GOS 的产量，在优化条件下达到 43%。mBgaD-D 和乳清粉的组合提高了牛奶的甜度，生产出了无添加糖、富含 GOS 的酸奶（GOSY）。GOSY 中的 GOS 含量为 4.1/100 g，提供了适当的甜度，并且产生了一种弹性和坚实的酸奶。GOSY 还增加了 24 小时粪便发酵过程中 spp.的数量。因此，用 mBgaD-D 和乳清粉强化酸奶可以提高其技术性能和健康益处。
