应用组合型四极杆-静电轨道阱质谱研究一种新合成二肽聚合物对糖肽的富集效果

Evaluating the Performance of a New Synthetic Dipeptide Polymer for Glycopeptide Enrichment on a Hybrid Quadrupole Orbitrap Mass Spectrometer

采用高分辨和高质量精度的组合型四极杆-静电轨道阱质谱(hybrid quadrupole orbitrap mass spectrometer)全面分析一种新合成的二肽聚合物对糖肽的富集效果,并由此建立一种评价材料对糖肽富集效果的方法。实验配制了一系列不同质量浓度比的牛胎球蛋白(bovine fetuin,糖蛋白)与牛血清蛋白(BSA,干扰蛋白)的混合干扰模型,先用胰蛋白酶酶解处理混合蛋白样品,再经合成的二肽聚合物富集样品中的糖肽。通过优化糖肽富集步骤和质谱分析条件,新合成的二肽聚合物可以从干扰蛋白浓度达目标蛋白100倍的样品中成功富集糖肽。与亲水性相互作用色谱(hydrophilic interaction liquid chromatography,HILIC)相比,该材料的富集性能远高于HILIC小柱。因此,该二肽聚合物能够提供更有效的糖肽富集,可为糖蛋白/糖肽的分离富集提供新途径。

Due to the significance of protein glycosylation in various biological processes,it is critical to characterize protein glycosylation accurately.In the past few decades,mass spectrometry(MS)has increasingly become a method to study protein glycosylation and made significant achievements in representing the roles of protein glycosylation in disease,such as cancer.In the general MS-based workflow,glycoproteins or glycopeptides usually require to be isolated from biological samples before MS analysis.Therefore,how to effectively enrich glycopeptides or glycoproteins from biological samples is crucial for successful characterization of protein glycosylation.Recently,a new material of dipeptide polymer(poly-PD@SiO2)was synthesized.In this study,the dipeptide polymer was used to enrich glycopeptides and a high resolution and high mass accuracy Hybrid Quadrupole Orbitrap mass spectrometer was utilized to evaluate the performance of enriching glycopeptides for this material.Firstly,fetuin and bovine serum albumin(BSA,as an interference protein)were mixed in different ratios to prepare a series of standard samples.Then,trypsin was used to digest samples.At last,glycopeptides were enriched by poly-PD@SiO2 before MS analysis.With the optimized enrichment procedures,poly-PD@SiO2 could successfully enrich glycopeptides from samples with resisting up to 100-fold BSA interference.However,for 10-fold BSA interference,samples enriched by hydrophilic interaction liquid chromatography(HILIC)were mainly non-glycopeptides.Therefore,it was demonstrated that poly-PD@SiO2 could provide more effective enrichment for glycopeptides in comparison with HILIC.