新型固定化铁离子亲和色谱整体柱的制备及评价

Preparation and Evaluation of Novel Iron Immobilized Metal Affinity Chromatography Monolith Column

以N-丙烯酰氧基琥珀酰亚胺为功能单体,制备了含有高活性基团的整体材料基质。以次氨基三乙酸为配体,通过固载Fe3+,发展了一种固定化Fe3+亲和色谱(Fe-IMAC)整体柱的制备方法。该整体柱不仅对磷酸化肽具有很好的选择性,而且富集容量大、回收率高、重现性好。此外,利用该整体柱实现了牛奶蛋白质酶解产物中磷酸化肽段的选择性富集。本实验研制的Fe-IMAC整体柱有望用于磷酸化蛋白质组研究。

A novel method to prepare monolithic Fe3＋ immobilized affinity chromatography column was developed, which included in situ polymerization of N-acryloxysuccinimide and ethylene dimethacrylate, nitilotriacetic acid activation, and Fe3＋ immobilization. The monolithic column showed high selectivity, high binding capacity, good recovery and good reproducibility for the enrichment of phosphopeptides. Such a column was successfully applied to the enrichment of phosphopeptides from the nonfat milk protein digests. All results demonstrate the promising of such a column in phosphoproteome analysis.