Solid-phase glycan isolation for glycomics analysis.
Solid-phase glycan isolation for glycomics analysis.
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DOI：
10.1002/prca.201200045
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发表时间：
2012-12
影响因子：
2
通讯作者：
Zhang, Hui
中科院分区：
文献类型：
--
作者：
Yang, Shuang;Zhang, Hui
糖基化是最重要的蛋白质翻译后修饰之一。蛋白质的生物活性会因其所含的聚糖而发生显著的变化。因此，在复杂的生物或临床样品中糖蛋白的聚糖的结构分析与聚糖与疾病的功能相关是至关重要的。通过HPLC-MS分析聚糖是分析聚糖结构和定量其在不同生物系统中的相对丰度的常用技术。依赖于MS的方法需要从可忽略的盐和其他污染物离子中分离聚糖，因为盐和离子可能干扰聚糖，导致聚糖电离不良。为了实现这些目标，已经开发了聚糖分离和净化方法，包括SPE、液相萃取、色谱和电泳。传统上，使用疏水性和亲水性柱的组合从蛋白质或肽中分离聚糖：蛋白质和肽保留在疏水性吸收剂上，而聚糖、盐和其他亲水性试剂作为流出物收集。然后通过亲水相互作用LC通过石墨-活性炭柱纯化流出物中的聚糖。然而，这些基于亲和力的方法的缺点是非特异性结合。因此，已经开发了通过酰肼或肟的化学方法，用于固相分离具有高特异性和产率的聚糖。结合高分辨率MS，特异性聚糖分离技术作为糖组学分析的有用工具提供了巨大的潜力。
Glycosylation is one of the most significant protein PTMs. The biological activities of proteins are dramatically changed by the glycans associated with them. Thus, structural analysis of the glycans of glycoproteins in complex biological or clinical samples is critical in correlation with the functions of glycans with diseases. Profiling of glycans by HPLC-MS is a commonly used technique in analyzing glycan structures and quantifying their relative abundance in different biological systems. Methods relied on MS require isolation of glycans from negligible salts and other contaminant ions since salts and ions may interfere with the glycans, resulting in poor glycan ionization. To accomplish those objectives, glycan isolation and clean-up methods including SPE, liquid-phase extraction, chromatography, and electrophoresis have been developed. Traditionally, glycans are isolated from proteins or peptides using a combination of hydrophobic and hydrophilic columns: proteins and peptides remain on hydrophobic absorbent while glycans, salts, and other hydrophilic reagents are collected as flowthrough. The glycans in the flowthrough are then purified through graphite-activated carbon column by hydrophilic interaction LC. Yet, the drawback in these affinity-based approaches is nonspecific binding. As a result, chemical methods by hydrazide or oxime have been developed for solid-phase isolation of glycans with high specificity and yield. Combined with high-resolution MS, specific glycan isolation techniques provide tremendous potentials as useful tools for glycomics analysis.
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