Institute of Biotechnology and Biochemical Engineering, Graz University of Technology, NAWI Graz, Petersgasse 12, A-8010, Graz, Austria.
Austrian Centre of Industrial Biotechnology, Krenngasse 37, A-8010, Graz, Austria.
Nat Commun. 2023 Nov 6;14(1):7123. doi: 10.1038/s41467-023-42750-0.
Biological degradation of natural product glycosides involves, alongside hydrolysis, β-elimination for glycosidic bond cleavage. Here, we discover an O-glycoside β-eliminase (OGE) from Agrobacterium tumefaciens that converts the C3-oxidized O-β-D-glucoside of phloretin (a plant-derived flavonoid) into the aglycone and the 2-hydroxy-3-keto-glycal elimination product. While unrelated in sequence, OGE is structurally homologous to, and shows effectively the same Mn active site as, the C-glycoside deglycosylating enzyme (CGE) from a human intestinal bacterium implicated in β-elimination of 3-keto C-β-D-glucosides. We show that CGE catalyzes β-elimination of 3-keto O- and C-β-D-glucosides while OGE is specific for the O-glycoside substrate. Substrate comparisons and mutagenesis for CGE uncover positioning of aglycone for protonic assistance by the enzyme as critically important for C-glycoside cleavage. Collectively, our study suggests convergent evolution of active site for β-elimination of 3-keto O-β-D-glucosides. C-Glycoside cleavage is a specialized feature of this active site which is elicited by substrate through finely tuned enzyme-aglycone interactions.
天然产物糖苷的生物降解除了水解外，还涉及糖苷键的β消除。在这里，我们从根癌农杆菌中发现了一种 O-糖苷β消除酶（OGE），它将植物来源的类黄酮根皮素的 C3-氧化的 O-β-D-葡萄糖苷转化为糖苷元和 2-羟基-3-酮糖消除产物。虽然 OGE 在序列上没有关系，但它在结构上与肠道细菌中的 C-糖苷去糖基化酶（CGE）同源，该酶参与 3-酮 C-β-D-葡萄糖苷的β消除。我们表明 CGE 催化 3-酮 O-和 C-β-D-葡萄糖苷的β消除，而 OGE 则特异性针对 O-糖苷底物。对 CGE 的底物比较和突变研究揭示了酶对糖苷元质子辅助的定位对于 C-糖苷裂解至关重要。总的来说，我们的研究表明，3-酮 O-β-D-葡萄糖苷β消除的活性位点发生了趋同进化。C-糖苷裂解是该活性位点的一个专门特征，通过底物与酶-糖苷元的精细相互作用来引发。
