基于凝集素-糖蛋白特异结合的特性筛选与鉴定空肠弯曲杆菌NCTC11168株糖蛋白

Screening and identification of glycoproteins from Campylobacter jejuni NCTC 11168 based on specific affinity between lectins and glycoproteins

摘要:

摘要:【目的】从空肠弯曲杆菌NCTC11168菌株中筛选糖蛋白。【方法】本实验基于凝集素-糖蛋白特异结合的特性，先用免疫印迹的方法确定空肠弯曲杆菌内糖蛋白能与凝集素SBA 特异结合，再用包被有凝集素SBA的磁珠捕获样品中的潜在糖蛋白，通过N-乙酰半乳糖胺竞争性洗脱及双向电泳分离，最后利用串联质谱鉴定捕获的糖蛋白。【结果】质谱分析共鉴定到22种空肠弯曲杆菌蛋白，其中至少5 种为已报道糖蛋白，包括Cj0633在内的17种为迄今为止未知的潜在糖蛋白。【结论】这种糖蛋白筛选策略可用于细菌糖蛋白的分离鉴定。

Abstract:

Abstract:［Objective］ To screen glycoproteins from Campylobacter jejuni NCTC 11168．［Methods］ We introduced a screening strategy based on specific affinity between lectins and glycoproteins． First，the specific affinity between Soybean Agglutinin (SBA) and glycoproteins in C． jejuni was confirmed by western blotting． Then，magnetic beads coating with Soybean Agglutinin (SBA) were used to capture the putative glycoproteins． Putative glycoproteins were competitively eluted by N-acetylgalactosamine and separated by two-dimensional gel electrophoresis． Spots in gels were further analyzed by mass spectrometry． ［Results］A total of 22 proteins were identified by mass spectrometric analysis，of which 5 have ever been identified to be glycoproteins． Others including Cj0633 have not been proved to be glycosylated so far．［Conclusion］The method could be used to screen glycoproteins in bacteria．