基于超滤反应器的蛋白质N-糖基化快速分析方法

Rapid N-glycosylation profiling of glycoprotein based on ultrafiltration-devices

随着生物科技的发展,蛋白质药物正越来越多地被应用在疾病诊疗的各个方面。但是蛋白质的匀质性,特别是蛋白质N-糖基化修饰成为限制蛋白质药物发展的瓶颈,因此蛋白质N-糖基化修饰的分析方法正逐渐成为蛋白质药物研发的关键技术。为了提高蛋白质N-糖基化修饰分析效率,本文建立了基于超滤反应器的N-糖分离纯化方法。新处理方法大幅提高了含N-乙酰基葡萄糖N-糖的提取效率（10%~20%）,避免了唾液酸结构的降解（20%~100%）,同时将处理时间缩短至30 min。通过与HPLC-HRMS分析方法联用,提高了样品分析效率。本文建立了高效、可靠,适合蛋白质药物的N-糖基化修饰分析方法。

With development of bio-technique, more and more proteins were applied as clinical approaches. However, the protein homogeneity, especially the N-glycosylation limited the further research and application of these protein drugs. The analysis method for N-glycans is believed to be critical in protein drugs development. To enhance the N-glycans isolation efficiency and accelerate the pretreatment, a new strategy was built on ultrafiltration-devices. New methods increased the isolation efficiency of N-glycans containing N-acetylglucosa mine with 10%-20%. The degrading of N-glycans containing sialic acids was also minimized with this method. 20%-100% more N-glycans with sialic acids were isolated. The pretreatment was finished within 30 min. Coupled with HPLC-HRMS, an effective and reliable strategy designed for protein drugs N-glycans analysis were developed.