One-step purification and immobilization of thermostable β-glucosidase on Na-Y zeolite based on the linker and its application in the efficient production of baohuoside I from icariin.

Baohuoside I, a minor flavonoid component of Herba Epimedii, has better bioactivities than its precursor compound icariin. In this work, we have fused the linker (4LP) to thermostable β-glucosidase (Tpebgl3) and successfully prepared the immobilized enzyme (4LP-Tpebgl3@Na-Y) to produce baohuoside I from icariin. The activity recovery and maximum load of 4LP-Tpebgl3@Na-Y were 95.4% and 50.3 mg/g, respectively. Moreover, it exhibited four-fold improved adsorption selectivity (80.5%) with respect to native enzyme after immobilization. The maximum activity of 4LP-Tpebgl3@Na-Y was exhibited at 85 °C, pH 5.0, and it retained>80% of its initial activity after incubation at 75 °C for 2 h . It showed enhanced tolerance of organic solvent and glucose as compared to free enzymes. Kcat/Km value for 4LP-Tpebgl3@Na-Y was 1616.0 s-1•mM-1, which was 61.0% higher than that of free enzyme. Under suitable conditions (75 °C, pH 5.0, 0.1 U/mL enzyme and 120 min), 2000 mg/L icariin was transformed into baohuoside I with a molar conversion of 97.6%. 4LP-Tpebgl3@Na-Y retained 85.2% of its original activity after 10 cycles of reuse and the 768.8 mg/L/h total productivity of baohuoside I was obtained. This is the first research on one-step purification and immobilization of thermostable β-glucosidase based on the linker and its application in the efficient production of baohuoside I from icariin.