Mass spectrometric analysis of the N-glycoproteome in statin-treated liver cells with two lectin-independent chemical enrichment methods.
Mass spectrometric analysis of the N-glycoproteome in statin-treated liver cells with two lectin-independent chemical enrichment methods.
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DOI：
10.1016/j.ijms.2017.05.010
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发表时间：
2018-06
影响因子：
1.8
通讯作者：
Wu R
中科院分区：
文献类型：
--
作者：
Xiao H;Hwang JE;Wu R
蛋白质N-糖基化是哺乳动物细胞生存所必需的，并且众所周知参与许多生物学过程。糖基化异常与癌症、传染病等人类疾病直接相关。蛋白质N-糖基化的全局分析将允许更好地理解蛋白质功能和细胞活性。质谱（MS）为基础的蛋白质组学提供了一个独特的机会，大规模的位点特异性表征蛋白质糖基化。由于生物样品的复杂性，有效的富集方法在MS分析之前至关重要。在这里，我们比较了两种凝集素独立的方法来丰富糖肽的蛋白质N-糖基化的MS的全球分析。第一个硼酸为基础的富集（BA）方法的好处，从硼酸和糖之间的普遍和可逆的相互作用;其他方法利用代谢标记和点击化学（MC），将化学处理到糖蛋白的未来亲和富集。我们全面比较了这两种方法在他汀类药物处理的肝细胞中糖蛋白的鉴定和定量方面的性能。基于目前的结果，BA方法在富集糖肽方面更通用，而MC方法高度富集细胞表面糖蛋白，并且定量结果似乎更动态，因为仅分析新合成的糖蛋白。此外，我们通过相应的亲本蛋白质比率标准化糖基化位点比率以反映真实的修饰变化。结合基于质谱的蛋白质组学，有效的富集方法将垂直推进蛋白质糖基化研究。
Protein N-glycosylation is essential for mammalian cell survival and is well-known to be involved in many biological processes. Aberrant glycosylation is directly related to human disease including cancer and infectious diseases. Global analysis of protein N-glycosylation will allow a better understanding of protein functions and cellular activities. Mass spectrometry (MS)-based proteomics provides a unique opportunity to site-specifically characterize protein glycosylation on a large scale. Due to the complexity of biological samples, effective enrichment methods are critical prior to MS analysis. Here, we compared two lectin-independent methods to enrich glycopeptides for the global analysis of protein N-glycosylation by MS. The first boronic acid-based enrichment (BA) method benefits from the universal and reversible interactions between boronic acid and sugars; the other method utilizes metabolic labeling and click chemistry (MC) to incorporate a chemical handle into glycoproteins for future affinity enrichment. We comprehensively compared the performance of the two methods in the identification and quantification of glycoproteins in statin-treated liver cells. Based on the current results, the BA method is more universal in enriching glycopeptides, while with the MC method, cell surface glycoproteins were highly enriched, and the quantification results appear to be more dynamic because only the newly-synthesized glycoproteins were analyzed. In addition, we normalized the glycosylation site ratios by the corresponding parent protein ratios to reflect the real modification changes. In combination with MS-based proteomics, effective enrichment methods will vertically advance protein glycosylation research.
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影响因子：
7.4
作者：
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通讯作者：
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影响因子：
64.5
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使用知情的搜索空间可最大程度地提高糖蛋白糖基化特定于位点特异性分配的信心。
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