Vinyl Sulfone Functionalization: A Feasible Approach for the Study of the Lectin–Carbohydrate Interactions

Carbohydrate-mediated molecular recognition is involved in many biological aspects such as cellular adhesion, immune response, blood coagulation, inflammation, and infection. Considering the crucial importance of such biological events in which proteins are normally involved, synthetic saccharide-based systems have emerged as powerful tools for the understanding of protein–carbohydrate interactions. As a new approach to create saccharide-based systems, a set of representative monosaccharides (d-mannose, d-glucose, N-acetyl-d-glucosamine, and l-fucose) and disaccharides (lactose, maltose, and melibiose) were derivatized at their anomeric carbon with a vinyl sulfone group spanned by an ethylthio linker. This vinyl sulfone functionalization is demonstrated to be a general strategy for the covalent linkage of a saccharide in mild conditions via Michael-type additions with the amine and thiol groups from functionalized supports and those naturally present in biomolecules. The introduction of the ethylthio linker between the biorecognizable element (i.e., saccharide) and the reactive group (i.e., vinyl sulfone) was found to preserve the functionality of the former. The capability of the vinyl sulfone saccharides for the study of lectin–carbohydrate interactions was demonstrated by (i) immobilizing them on both amine-functionalized supports (glass slides and microwell plates) and polylysine-coated glass slides to create sugar arrays that selectively bind lectins (ii) coupling to model proteins to yield neoglycoproteins that are recognized by lectins and (iii) using vinyl sulfone saccharides as tags to allow the detection of the labeled biomolecule by HRP-lectins. The above results were further put tothe test with a real case: detection of carbohydrate binding proteins present in rice (Oryza sativa).碳水化合物介导的分子识别参与诸多生物学过程，诸如细胞黏附、免疫应答、血液凝固、炎症反应与感染过程。鉴于这类通常由蛋白质参与的生物学事件至关重要，基于合成糖类的体系已成为解析蛋白质-糖类相互作用的有力工具。作为构建糖类体系的全新策略，研究人员对一系列代表性单糖（D-甘露糖、D-葡萄糖、N-乙酰-D-氨基葡萄糖以及L-岩藻糖）和二糖（乳糖、麦芽糖、蜜二糖）的端基碳进行了衍生化修饰，引入了由乙硫基连接臂连接的乙烯砜基团。实验证明，这种乙烯砜官能化策略是一种通用方法，可在温和条件下通过迈克尔加成反应（Michael-type additions），将糖类共价连接至功能化载体表面的氨基、巯基，以及天然存在于生物分子中的氨基与巯基之上。研究发现，在可生物识别元件（即糖类）与反应活性基团（即乙烯砜）之间引入乙硫基连接臂，能够保留前者的生物功能。乙烯砜修饰糖类用于研究凝集素（lectin）-糖类相互作用的能力，通过以下三方面得到验证：(i) 将其固定于氨基功能化载体（玻璃玻片与微孔板）以及聚赖氨酸包被的玻璃玻片上，制备可选择性结合凝集素的糖芯片阵列（sugar arrays）；(ii) 将其与模式蛋白偶联，制备可被凝集素识别的新糖蛋白（neoglycoproteins）；(iii) 以乙烯砜修饰糖类作为标记标签，实现通过辣根过氧化物酶（horseradish peroxidase, HRP）偶联凝集素对标记生物分子的检测。上述研究结果还通过一个实际案例得到了进一步验证：检测水稻（Oryza sativa）中存在的糖类结合蛋白。