Department of Chemistry, University of California, Davis, CA 95616, USA.
Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, China.
Molecules. 2021 Oct 14;26(20):6194. doi: 10.3390/molecules26206194.
Carbohydrate-protein conjugates have diverse applications. They have been used clinically as vaccines against bacterial infection and have been developed for high-throughput assays to elucidate the ligand specificities of glycan-binding proteins (GBPs) and antibodies. Here, we report an effective process that combines highly efficient chemoenzymatic synthesis of carbohydrates, production of carbohydrate-bovine serum albumin (glycan-BSA) conjugates using a squarate linker, and convenient immobilization of the resulting neoglycoproteins on carboxylate-coated fluorescent magnetic beads for the development of a suspension multiplex array platform. A glycan-BSA-bead array containing BSA and 50 glycan-BSA conjugates with tuned glycan valency was generated. The binding profiles of six plant lectins with binding preference towards Gal and/or GalNAc, as well as human galectin-3 and galectin-8, were readily obtained. Our results provide useful information to understand the multivalent glycan-binding properties of human galectins. The neoglycoprotein-immobilized fluorescent magnetic bead suspension multiplex array is a robust and flexible platform for rapid analysis of glycan and GBP interactions and will find broad applications.
碳水化合物-蛋白质偶联物具有多种应用。它们已被临床用于针对细菌感染的疫苗，并被开发用于高通量测定法，以阐明糖结合蛋白 (GBP) 和抗体的配体特异性。在这里，我们报告了一种有效的方法，该方法结合了高效的化学酶合成碳水化合物、使用 squarate 接头生产碳水化合物-牛血清白蛋白 (glycan-BSA) 偶联物，以及方便地将所得的糖基化蛋白固定在羧酸盐涂覆的荧光磁性珠上，用于开发悬浮液多重分析平台。生成了包含 BSA 和 50 种糖基化 BSA 缀合物的糖基化 BSA-珠阵列，其中糖基化 BSA 缀合物的糖基化价得到了调节。容易获得六种植物凝集素与 Gal 和/或 GalNAc 的结合偏好，以及人半乳糖凝集素-3 和半乳糖凝集素-8 的结合特征。我们的结果提供了有用的信息，有助于理解人半乳糖凝集素的多价糖结合特性。固定有糖基化蛋白的荧光磁性珠悬浮液多重分析平台是一种快速分析聚糖和 GBP 相互作用的强大而灵活的平台，具有广泛的应用前景。
