Biomedical Engineering Department, Northwestern University, Evanston, IL 60208, USA.
J Sep Sci. 2012 Jun;35(12):1514-20. doi: 10.1002/jssc.201200073.
Inspired by the catechol-rich adhesive proteins of the mussel foot, we report a simple and versatile aqueous approach for the immobilization of trypsin onto silica and titania monolithic supports. The method involves in-situ coating of the monolithic substrates with a catechol-containing biomimetic polymer (polydopamine) derived from the polymerization of dopamine under alkaline pH, followed by conjugation of trypsin to the polydopamine polymer coating. The trypsin immobilization efficiency onto the monolithic materials was investigated as a function of different preparation parameters such as dopamine concentration and coating time. The enzymatic activity of the immobilized trypsin reactors was evaluated, and mass spectrometry based proteomic analysis was demonstrated by digestion of a model protein. The method presented in this manuscript has broad potential for immobilization of trypsin and other enzymes onto a wide variety of monolithic supports, due to the ability of polydopamine to act as a primer for covalent immobilization of proteins.
受贻贝足部富含儿茶酚的黏附蛋白的启发，我们报告了一种简单而通用的水相方法，用于将胰蛋白酶固定在二氧化硅和二氧化钛整体式载体上。该方法包括通过在碱性 pH 下聚合多巴胺，原位涂覆含有儿茶酚的仿生聚合物（聚多巴胺），然后将胰蛋白酶结合到聚多巴胺聚合物涂层上，来实现整体式基质的原位涂覆。研究了不同制备参数（例如多巴胺浓度和涂层时间）对整体式材料上胰蛋白酶固定效率的影响。评估了固定化胰蛋白酶反应器的酶活性，并通过消化模型蛋白进行了基于质谱的蛋白质组学分析。由于聚多巴胺能够作为蛋白质共价固定的底漆，因此本文提出的方法具有将胰蛋白酶和其他酶固定在各种整体式载体上的广泛潜力。
