Kaya Masahiko, Ito Junji, Kotaka Atsushi, Matsumura Kengo, Bando Hiroki, Sahara Hiroshi, Ogino Chiaki, Shibasaki Seiji, Kuroda Kouichi, Ueda Mitsuyoshi, Kondo Akihiko, Hata Yoji
Research Institute, Gekkeikan Sake Co Ltd, Kyoto, Japan.
Appl Microbiol Biotechnol. 2008 May;79(1):51-60. doi: 10.1007/s00253-008-1393-6. Epub 2008 Mar 14.
For efficient production of isoflavone aglycones from soybean isoflavones, we isolated three novel types of beta-glucosidase (BGL1, BGL3, and BGL5) from the filamentous fungi Aspergillus oryzae. Three enzymes were independently displayed on the cell surface of a yeast Saccharomyces cerevisiae as a fusion protein with alpha-agglutinin. Three beta-glucosidase-displaying yeast strains hydrolyzed isoflavone glycosides efficiently but exhibited different substrate specificities. Among these beta-glucosidases, BGL1 exhibited the highest activity and also broad substrate specificity to isoflavone glycosides. Although glucose released from isoflavone glycosides are generally known to inhibit beta-glucosidase, the residual ratio of isoflavone glycosides in the reaction mixture with BGL1-displaying yeast strain (Sc-BGL1) reached approximately 6.2%, and the glucose concentration in the reaction mixture was maintained at lower level. This result indicated that Sc-BGL1 assimilated the glucose before they inhibited the hydrolysis reaction, and efficient production of isoflavone aglycones was achieved by engineered yeast cells displaying beta-glucosidase.
为了从大豆异黄酮高效生产异黄酮苷元，我们从丝状真菌米曲霉中分离出三种新型β-葡萄糖苷酶（BGL1、BGL3和BGL5）。这三种酶作为与α-凝集素的融合蛋白独立展示在酿酒酵母的细胞表面。三种展示β-葡萄糖苷酶的酵母菌株能高效水解异黄酮苷，但表现出不同的底物特异性。在这些β-葡萄糖苷酶中，BGL1活性最高，对异黄酮苷也具有广泛的底物特异性。虽然通常认为从异黄酮苷释放的葡萄糖会抑制β-葡萄糖苷酶，但在含有展示BGL1的酵母菌株（Sc-BGL1）的反应混合物中，异黄酮苷的残留率达到约6.2%，且反应混合物中的葡萄糖浓度保持在较低水平。这一结果表明，Sc-BGL1在葡萄糖抑制水解反应之前就将其同化，通过展示β-葡萄糖苷酶的工程酵母细胞实现了异黄酮苷元的高效生产。
