Department of Applied Biology and Chemical Technology, Central Laboratory of the Institute of Molecular Technology for Drug Discovery and Synthesis, The Hong Kong Polytechnic University, Hung Hom, Kowloon, Hong Kong, PR China.
Carbohydr Res. 2008 Nov 24;343(17):2932-8. doi: 10.1016/j.carres.2008.08.021. Epub 2008 Aug 28.
Carbohydrate biosensors, including carbohydrate arrays, are attracting increased attention for the comprehensive and high-throughput investigation of protein-carbohydrate interactions. Here, we describe an effective approach to fabricating a robust microplate-based carbohydrate array capable of probing protein binding and screening for inhibitors in a high-throughout manner. This approach involves the derivatization of carbohydrates with a trityl group through an alkyl linker and the immobilization of the trityl-derivatized carbohydrates (mannose and maltose) onto microplates noncovalently to construct carbohydrate arrays. The trityl carbohydrate derivative has very good immobilization efficiency for polystyrene microplates and strong resistance to aqueous washing. The carbohydrate arrays can probe the interactions with the lectin Concanavalin A and screen this protein for the well-known inhibitors methyl alpha-D-mannopyranoside and methyl alpha-D-glucopyranoside in a high-throughput manner. The method described in this paper represents a convenient way of fabricating robust noncovalent carbohydrate arrays on microplates and offers a convenient platform for high-throughput drug screening.
碳水化合物生物传感器,包括碳水化合物微阵列,由于能够高通量全面地研究蛋白质-碳水化合物相互作用而受到越来越多的关注。在这里,我们描述了一种有效的方法来制备稳健的基于微板的碳水化合物微阵列,能够以高通量的方式探测蛋白质结合并筛选抑制剂。该方法涉及通过烷基 linker 将碳水化合物衍生化为三苯甲基基团,并将三苯甲基衍生化的碳水化合物(甘露糖和麦芽糖)非共价固定在微板上以构建碳水化合物微阵列。三苯甲基碳水化合物衍生物对聚苯乙烯微板具有非常好的固定效率,并且对水性洗涤具有很强的抵抗力。碳水化合物微阵列可以探测与凝集素 Concanavalin A 的相互作用,并以高通量的方式筛选该蛋白的已知抑制剂甲基α-D-吡喃甘露糖苷和甲基α-D-吡喃葡萄糖苷。本文所述的方法提供了一种在微板上制备稳健的非共价碳水化合物微阵列的简便方法,并为高通量药物筛选提供了一个方便的平台。