Programa de Glicobiologia, Instituto de Bioquímica Médica Leopoldo de Meis, and Laboratório de Tecido Conjuntivo, Hospital Universitário Clementino Fraga Filho, Universidade Federal do Rio de Janeiro, Rio de Janeiro, RJ 21941-913, Brazil.
Glycobiology. 2017 Jul 1;27(7):625-634. doi: 10.1093/glycob/cwx031.
Fucosylated chondroitin sulfate (FCS) from sea cucumbers is composed of a chondroitin sulfate (CS) central core and branches of sulfated fucose. The structure of this complex glycosaminoglycan is usually investigated via nuclear magnetic resonance (NMR) analyses of the intact molecule, ergo through a top-down approach, which often yield spectra with intricate sets of signals. Here we employed a bottom-up approach to analyze the FCSs from the sea cucumbers Isostichopus badionotus and Ludwigothurea grisea from their basic constituents, viz. CS cores and sulfated fucose branches, obtained via systematic fragmentation through mild acid hydrolysis. Oligosaccharides derived from the central CS core were analyzed via NMR spectroscopy and the disaccharides produced using chondroitin sulfate lyase via SAX-HPLC. The CS cores from the two species were similar, showing only slight differences in the proportions of 4- or 6-monosulfated and 4,6-disulfated β-d-GalNAc. Sulfated fucose units released from the FCSs were analyzed via NMR and ESI-HRMS spectroscopies. The fucose units from each species presented extensive qualitative differences, but quantitative assessments of these units were hindered, mostly because of their extensive desulfation during the hydrolysis. The bottom-up analysis performed here has proved useful to explore the structure of FCS through a sum-of-the-parts approach in a qualitative manner. We further demonstrate that under specific acidification conditions particular fucose branches can be removed preferentially from FCS. Preparation of derivatives enriched with particular fucose branches could be useful for studies on "structure vs. biological function" of FCS.
海参岩黄芪糖胺聚糖（FCS）由核心的硫酸软骨素（CS）和分支的硫酸岩藻糖组成。这种复杂糖胺聚糖的结构通常通过完整分子的核磁共振（NMR）分析来研究，也就是自上而下的方法，该方法通常会产生具有复杂信号集的光谱。在这里，我们采用自下而上的方法，从海参黄海胆和灰海参的基本成分（即 CS 核心和硫酸岩藻糖分支）来分析 FCS，这些基本成分是通过温和酸水解进行系统断裂得到的。通过 NMR 光谱分析从中获得的 CS 核心寡糖，并使用软骨素酶通过 SAX-HPLC 分析二糖。两种海参的 CS 核心相似，只是 4-或 6-单硫酸化和 4,6-二硫酸化β-d-GalNAc 的比例略有不同。通过 NMR 和 ESI-HRMS 光谱分析从 FCS 中释放的硫酸岩藻糖单元。每个物种的岩藻糖单元呈现出广泛的定性差异，但对这些单元的定量评估受到阻碍，主要是因为在水解过程中它们广泛脱硫酸化。这里进行的自下而上的分析已被证明可通过部分总和方法以定性方式探究 FCS 的结构。我们进一步证明，在特定酸化条件下，可以优先从 FCS 中去除特定的岩藻糖分支。制备富含特定岩藻糖分支的衍生物可能有助于研究 FCS 的“结构与生物学功能”。
