Immobilization of pectin fragments on solid supports: Novel coupling by thiazolidine formation
Immobilization of pectin fragments on solid supports: Novel coupling by thiazolidine formation
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DOI：
10.1021/bc0155364
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发表时间：
2002-03-01
影响因子：
4.7
通讯作者：
Jensen, KJ
中科院分区：
文献类型：
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作者：
Guillaumie, F;Thomas, ORT;Jensen, KJ
作为固相和序列分析以及果胶精细结构研究的先决条件，我们开发了定向和化学选择性方法，将模型果胶片段偶联到固体支持物上。选择聚乙二醇聚丙烯酰胺 (PEGA) 树脂是因为它们在包括水在内的多种溶剂中具有出色的溶胀性能，并且易于与酶接触。对氨基封端的 PEGA 树脂进行适当的衍生化后，α-D-半乳糖醛酸 (GalA) 的低聚物（直至三聚体）通过其还原端锚定到载体上。除了还原胺化之外，这些策略还包括形成肟键、糖基酰肼和焦谷氨酰环。此外，我们开发了一种基于噻唑烷环形成的新固定方法。所有方法都被证明是有效的，并且不需要在偶联前对 GalA 寡聚物进行修饰。此外，支持物的衍生化需要非常温和的条件和很少的步骤。考虑到形成的键的稳定性、与水性溶剂的相容性、所需的步骤数以及应用于较大果胶片段的潜力，通过噻唑烷环固定和形成肟键是优选的方法。通过插入二硫键进一步发展了噻唑烷和焦谷氨酰锚定，这允许在温和的选择性条件下释放糖。
As a prerequisite to solid-phase and sequence analyses and for the study of the fine structure of pectin, we have developed oriented and chemoselective methodologies to couple model pectin fragments onto a solid support. Polyethylene glycol polyacrylamide (PEGA) resins were selected due to their excellent swelling properties in a wide range of solvents, including water, and their easy accessibility to enzymes. Following appropriate derivatization of amino-terminated PEGA resins, oligomers of alpha-D-galacturonic acid (GalA), up to the trimer, were anchored to the support through their reducing end. In addition to reductive amination, the strategies included the formation of an oxime bond, a glycosyl hydrazide, and a pyroglutamyl ring. Further, we developed a new immobilization approach based on the formation of a thiazolidine ring. All methods proved efficient and did not require modification of the GalA oligomers prior to coupling. In addition, very mild conditions and few steps for derivatization of the support were required. Immobilization by thiazolidine ring and oxime bond formation were the preferred methods, given the stability of the linkages formed, their compatibility with aqueous solvents, the few number of steps required, and their potential for application to larger pectin fragments. Thiazolidine and pyroglutamyl anchoring were developed further by the insertion of a disulfide bond which allowed release of the saccharides under mild, selective conditions.