Improved methods to characterize the length and quantity of highly unstable PolySialic acids subject category: (Carbohydrates, chromatographic techniques).
Improved methods to characterize the length and quantity of highly unstable PolySialic acids subject category: (Carbohydrates, chromatographic techniques).
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DOI：
10.1016/j.ab.2021.114426
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发表时间：
2021-12-15
影响因子：
2.9
通讯作者：
中科院分区：
文献类型：
--
作者：
聚唾液酸（polySia）是α2-8-连接唾液酸的线性均聚物，在哺乳动物脑发育的早期阶段高度表达，并调节多种细胞功能。虽然聚合度（DP）可以影响这些功能，但是由于聚合物的不稳定性，目前可用的方法不能准确地表征该参数。我们开发了两种改进的方法来表征生物样品中的DP和总polySia含量。具有暴露的还原末端的PolySia链可以用DMB衍生化用于随后的HPLC分析。然而，应用于聚Sia-糖蛋白的生物样品需要从底层聚糖释放聚Sia链，这在不同时部分水解聚Sia链的α2-8-键的情况下难以实现，从而影响其准确表征。我们报告了一种保护长polySia链的内部α2- 8 sia键的方法，使用先前已知的酯化条件产生稳定的聚内酯结构。这种聚内酯化的分子在酸水解释放和酸性DMB衍生化过程中更稳定。此外，我们使用高度特异性的Endoneuraminidase-NF酶来区分聚唾液酸和其他唾液酸，并开发了一种精确测量生物样品中polySia总含量的方法。这两种方法提供了改进的polySia的定量和表征。
Polysialic acid (polySia) is a linear homopolymer of α2-8-linked sialic acids that is highly expressed during early stages of mammalian brain development and modulates a multitude of cellular functions. While degree of polymerization (DP) can affect such functions, currently available methods do not accurately characterize this parameter, because of the instability of the polymer. We developed two improved methods to characterize the DP and total polySia content in biological samples. PolySia chains with exposed reducing termini can be derivatized with DMB for subsequent HPLC analysis. However, application to biological samples of polySia-glycoproteins requires release of polySia chains from the underlying glycan, which is difficult to achieve without concurrent partial hydrolysis of the α2-8-linkages of the polySia chain, affecting its accurate characterization. We report an approach to protect internal α2-8sia linkages of long polySia chains, using previously known esterification conditions that generate stable polylactone structures. Such polylactonized molecules are more stable during acid hydrolysis release and acidic DMB derivatization. Additionally, we used the highly specific Endoneuraminidase-NF enzyme to discriminate polysialic acid and other sialic acid and developed an approach to precisely measure the total content of polySia in a biological sample. These two methods provide improved quantification and characterization of polySia.
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影响因子：
3.4
作者：
KATCHALSKY, A;SPITNIK, P
通讯作者：
SPITNIK, P
影响因子：
2.9
作者：
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Inoue, Y
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GOEBEL, WF
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Inoue, Y
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作者：
Galuska, Sebastian P.;Geyer, Rudolf;Geyer, Hildegard
通讯作者：
Geyer, Hildegard