Catalytic properties characterization and degradation mode elucidation of a polyG-specific alginate lyase OUC-FaAly7.

Polymerized guluronates (polyG)-specific alginate lyase with lower polymerized mannuronates (polyM)-degrading activity, superior stability, and clear action mode is a powerful biotechnology tool for the preparation of AOSs rich in M blocks. In this study, we expressed and characterized a polyG-specific alginate lyase OUC-FaAly7 from Formosa agariphila KMM3901. OUC-FaAly7 belonging to polysaccharide lyase (PL) family 7 had highest activity (2743.7 ± 20.3 U/µmol) at 45 °C and pH 6.0. Surprisingly, its specific activity against polyG reached 8560.2 ± 76.7 U/µmol, whereas its polyM-degrading activity was nearly 0 within 10 min reaction. Suggesting that OUC-FaAly7 was a strict polyG-specific alginate lyase. Importantly, OUC-FaAly7 showed a wide range of temperature adaptations and remarkable temperature and pH stability. Its relative activity between 20 °C and 45 °C reached >90 % of the maximum activity. The minimum identifiable substrate of OUC-FaAly7 was guluronate tetrasaccharide (G4). Action process and mode showed that it was a novel alginate lyase digesting guluronate hexaose (G6), guluronate heptaose (G7), and polymerized guluronates, with the preferential generation of unsaturated guluronate pentasaccharide (UG5), although which could be further degraded into unsaturated guluronate disaccharide (UG3) and trisaccharide (UG2). This study contributes to illustrating the catalytic properties, substrate recognition, and action mode of novel polyG-specific alginate lyases.

High-level and reusable preparation of sulforaphane by yeast cells expressing myrosinase.

Myrosinase is a key tool for the fast and efficient preparation of sulforaphane which is one of the prominent natural ingredients found in brassicaceous vegetables. Here, the glucoraphanin-hydrolyzing activity of a Yarrowia lipolytica 20-8 harboring myrosinase reached 73.28 U/g dry cell weight, indicating that it had a potential application in sulforaphane preparation from glucoraphanin. An efficient and reusable process for sulforaphane preparation via myrosinase produced by Y. lipolytica 20-8 was constructed. In detail, as high as 10.32 mg sulforaphane could be produced from 1 g broccoli seed under the reaction of 40 U yeast whole-cell catalyst within 15 min with the conversion efficiency of 99.86%. Moreover, when the yeast whole-cell catalyst was reused 7 and 10 times, as high as 92.53% and 87.56% of sulforaphene yield of the initial level could be retained, respectively. Therefore, this yeast whole-cell is a potent biocatalyst for the efficient and reusable preparation of sulforaphane.