Departamento de Microbiología y Bioquímica de Productos Lácteos, Instituto de Productos Lácteos de Asturias (IPLA-CSIC), Paseo Río Linares, s/n, 33300 Villaviciosa, Asturias, Spain.
Departamento de Microbiología y Bioquímica de Productos Lácteos, Instituto de Productos Lácteos de Asturias (IPLA-CSIC), Paseo Río Linares, s/n, 33300 Villaviciosa, Asturias, Spain.
Food Res Int. 2017 Oct;100(Pt 1):522-528. doi: 10.1016/j.foodres.2017.07.024. Epub 2017 Jul 21.
Bifidobacterium pseudocatenulatum IPLA 36007 acts on isoflavone glycosides, releasing their corresponding aglycones. This strain-specific activity might be a key step in making isoflavones bioavailable and harnessing their oestrogenic activity. To investigate the molecular mechanisms involved in this activity, four glycosyl hydrolase-encoding genes in the IPLA 36007 genome (AW18_01575, AW18_09810, AW18_08145, and AW18_08090) were selected, synthesized with heterologous promoter and terminator signals (r-β-gluA, r-β-gluB, r-β-gluD and r-β-gluE, respectively), cloned into Escherichia coli, overexpressed as His-tagged proteins, and the enzymes purified and characterized. All four enzymes - GluA, GluB, GluD and GluE - proved to have β-glucosidase activity and deglycosylated (although at different rates) the isoflavone glycosides daidzin and genistin, releasing the aglycone moieties daidzein and genistein, respectively. GluD and GluE were also shown to hydrolyse β-glucosyl disaccharides such as cellobiose and gentiobiose, while GluA and GluB did not. Differences in activity were recorded for all four β-glucosidases at different pHs and temperatures under otherwise similar assay conditions, suggesting they have complementary activities under different environmental conditions. Two of the recombinant genes, r-β-gluA, and r-β-gluD, were cloned and expressed in the model lactic acid bacterium Lactococcus lactis, suggesting starter and probiotic organisms could be endowed with β-glucosidase activity. B. pseudocatenulatum IPLA 36007 contains additional β-glucosidases to those studied in this work, indicating a high level of redundancy for this enzymatic activity. Knowledge of glycoside-degrading enzymes should facilitate the development of novel, more effective or more selective prebiotics or functional foods for the promotion of bifidobacterial numbers in the human gut. It might also be of interest in the development of novel probiotics with specific health-promoting activities.
植物双歧杆菌 IPLA 36007 作用于异黄酮糖苷，释放出相应的苷元。这种菌株特异性的活性可能是使异黄酮具有生物利用度并利用其雌激素活性的关键步骤。为了研究涉及这种活性的分子机制，在 IPLA 36007 基因组中选择了四个糖苷水解酶编码基因（AW18_01575、AW18_09810、AW18_08145 和 AW18_08090），分别用异源启动子和终止子信号（r-β-gluA、r-β-gluB、r-β-gluD 和 r-β-gluE）合成，克隆到大肠杆菌中，作为 His 标记蛋白过表达，并对酶进行纯化和表征。四种酶（GluA、GluB、GluD 和 GluE）均具有β-葡萄糖苷酶活性，并使异黄酮糖苷大豆苷元和染料木苷发生去糖基化（尽管速率不同），分别释放出苷元大豆黄素和染料木黄酮。还发现 GluD 和 GluE 水解β-葡萄糖苷二糖，如纤维二糖和龙胆二糖，而 GluA 和 GluB 则不水解。在不同的 pH 值和温度下，在其他相似的测定条件下，记录了所有四种β-葡萄糖苷酶的活性差异，这表明它们在不同的环境条件下具有互补的活性。在模式乳杆菌 Lactococcus lactis 中克隆并表达了两种重组基因 r-β-gluA 和 r-β-gluD，这表明启动子和益生菌可以具有β-葡萄糖苷酶活性。B. pseudocatenulatum IPLA 36007 含有比本研究中研究的更多的β-葡萄糖苷酶，表明这种酶活性具有高度的冗余性。糖苷降解酶的知识应该有助于开发新型、更有效或更具选择性的益生元或功能性食品，以促进双歧杆菌在人类肠道中的数量。它可能也有助于开发具有特定健康促进活性的新型益生菌。
