Highly Efficient Analysis of Glycoprotein Sialylation in Human Serum by Simultaneous Quantification of Glycosites and Site-Specific Glycoforms
Highly Efficient Analysis of Glycoprotein Sialylation in Human Serum by Simultaneous Quantification of Glycosites and Site-Specific Glycoforms
复制标题
通过同时定量糖位点和位点特异性糖型来高效分析人血清中的糖蛋白唾液酸化
DOI：
10.1021/acs.jproteome.9b00332
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发表时间：
2019-09-01
影响因子：
4.4
通讯作者：
Ye, Mingliang
中科院分区：
文献类型：
--
作者：
Qin, Hongqiang;Dong, Xuefang;Ye, Mingliang
糖蛋白的异常唾液酸化与许多恶性疾病密切相关，唾液酸化分析对揭示这些疾病的状态具有巨大的潜力。然而，由于蛋白质糖基化的高度微观异质性以及唾液酸化糖肽（SGPs）的低丰度，唾液酸化的深入分析仍然具有挑战性。在此，通过采用SGP富集方法，在糖位点和位点特异性糖型的水平上制作了用于糖蛋白唾液酸化的详细表征的综合策略。该策略能够识别多达380个糖位点，以及414个完整的糖肽，对应于383个位点特异性糖型，仅从最初的6 μ L血清样品，表明高灵敏度的糖蛋白唾液酸化的详细分析的方法。该策略被进一步用于肝细胞癌患者和对照样品之间糖蛋白唾液酸化的差异分析，同时导致344个糖位点和405个位点特异性糖型的定量。其中，43个糖位点和55个位点特异性糖型分别在糖位点和位点特异性糖型水平上发生了显著变化。有趣的是，几个糖型连接到同一个糖位点上，发现有不同的变化趋势。这一策略被证明是一个强有力的工具，揭示糖蛋白唾液酸化的宏观和微观异质性的细微差异。
Aberrant sialylation of glycoproteins is closely related to many malignant diseases, and analysis of sialylation has great potential to reveal the status of these diseases. However, in-depth analysis of sialylation is still challenging because of the high microheterogeneity of protein glycosylation, as well as the low abundance of sialylated glycopeptides (SGPs). Herein, an integrated strategy was fabricated for the detailed characterization of glycoprotein sialylation on the levels of glycosites and site-specific glycoforms by employing the SGP enrichment method. This strategy enabled the identification of up to 380 glycosites, as well as 414 intact glycopeptides corresponding to 383 site-specific glycoforms from only initial 6 mu L serum samples, indicating the high sensitivity of the method for the detailed analysis of glycoprotein sialylation. This strategy was further employed to the differential analysis of glycoprotein sialylation between hepatocellular carcinoma patients and control samples, leading to the quantification of 344 glycosites and 405 site-specific glycoforms, simultaneously. Among these, 43 glycosites and 55 site-specific glycoforms were found to have significant change on the glycosite and site-specific glycoform levels, respectively. Interestingly, several glycoforms attached onto the same glycosite were found with different change tendencies. This strategy was demonstrated to be a powerful tool to reveal subtle differences of the macro- and microheterogeneity of glycoprotein sialylation.