Highly Efficient Biotransformation of Astragaloside IV to Cycloastragenol by Sugar-Stimulated β-Glucosidase and β-Xylosidase from Dictyoglomus thermophilum

beta-Glucosidases and beta-xylosidases are two categories of enzymes that could cleave out non-reducing, terminal beta-D-glucosyl and beta-D-xylosyl residues with release of D-glucose and D-xylose, respectively. In this paper, two functional beta-glucosidase Dth3 and beta-xylosidase Xln-DT from Dictyoglomus thermophilum were heterologously expressed in E.coli BL21 (DE3). Dth3 and Xln-DT were relatively stable at 75 degrees C and were tolerant or even stimulated by glucose and xylose. Dth3 was highly tolerant to glucose with a K-i value of approximately 3 M. Meanwhile, it was not affected by xylose in high concentration. The activity of Xln-DT was stimulated 2.13-fold by 1 M glucose and 1.29-fold by 0.3 M xylose, respectively. Furthermore, the beta-glucosidase Dth3 and beta-xylosidase Xln-DT showed excellent selectivity to cleave the outer C-6 and C-3 sugar moieties of ASI, which established an effective and green method to produce the more pharmacologically active CAG, an exclusive telomerase activator. We measured temperature, pH and dosage of enzyme using a single-factor experiment in ASI biotransformation. After optimization, the optimal reaction conditions were as follows: 75 degrees C, pH 5.5, 1 U of Dth3 and 0.2 U of Xln- DT, respectively. Under the optimized conditions, 1 g/l ASI was transformed into 0.63 g/l CAG with a corresponding molar conversion of 94.5% within 3 h. This is the first report to use the purified thermostable and sugar-tolerant enzymes from Dictyoglomus thermophilum to hydrolyze ASI synergistically, which provides a specific, environment-friendly and cost-effective way to produce CAG.