The elucidation of glycans biological function is essential to understand their role in biological processes, both normal and pathological. Immobilized glycoenzymes are excellent tools for this purpose as they can selectively release glycans from glycoproteins without altering their backbone. They can be easily removed from the reaction mixture avoiding their interference in subsequent experiments. Here, we describe the immobilization of peptide-N-glycosidase F (PNGase F) onto silica magnetic nanoparticles with immobilization yields of 86% and activity yields of 12%. Immobilized PNGase F showed higher thermal stability than its soluble counterpart, and could be reused for at least seven deglycosylation cycles. It was efficient in the deglycosylation of several glycoproteins (ribonuclease B, bovine fetuin, and ovalbumin) and a protein lysate from the parasite Fasciola hepatica under native conditions, with similar performance to that of the soluble enzyme. Successful deglycosylation was evidenced by a decrease in specific lectin recognition of the glycoproteins (40%–80%). Moreover, deglycosylated F. hepatica lysate allowed us to confirm the role of parasite N-glycans in the inhibition of the lipopolysaccharide-induced maturation of dendritic cells. Immobilized PNGase F probed to be a robust biotechnological tool for deglycosylation of glycoproteins and complex biological samples under native conditions.

中文翻译：

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将肽-N-糖苷酶 F 固定在磁性纳米粒子上：天然条件下蛋白质去糖基化的生物技术工具

聚糖生物学功能的阐明对于了解它们在正常和病理生物学过程中的作用至关重要。固定化糖酶是用于此目的的极好工具，因为它们可以选择性地从糖蛋白中释放聚糖而不改变其骨架。它们可以很容易地从反应混合物中去除，避免它们对后续实验的干扰。在这里，我们描述了将肽-N-糖苷酶 F (PNGase F) 固定在二氧化硅磁性纳米颗粒上，固定产率为 86%，活性产率为 12%。固定化 PNGase F 比其可溶性对应物表现出更高的热稳定性，并且可以重复使用至少七个去糖基化循环。它对几种糖蛋白（核糖核酸酶 B、牛胎球蛋白、天然条件下的肝片状菌，与可溶性酶具有相似的性能。糖蛋白的特异性凝集素识别减少（40%–80%）证明了成功的去糖基化。此外，去糖基化肝梭菌裂解物使我们能够确认寄生虫 N-聚糖在抑制脂多糖诱导的树突状细胞成熟中的作用。固定化 PNGase F 被认为是一种强大的生物技术工具，用于在天然条件下对糖蛋白和复杂生物样品进行去糖基化。