Jennison Amy V, Raqib Rubhana, Verma Naresh K
School of Biochemistry and Molecular Biology, Faculty of Science, The Australian National University, Canberra, ACT 0200, Australia.
World J Gastroenterol. 2006 Nov 7;12(41):6683-8. doi: 10.3748/wjg.v12.i41.6683.
To profile the immunogenic proteins of Shigella flexneri (S. flexneri) expressed during human infection using a proteomic approach.
Soluble and membrane protein extractions of S. flexneri 2457T were separated by two-dimensional gel electrophoresis (2-DE). Proteins were transferred to PVDF membrane and immunoblotted with sera from shigellosis patients. Reactive protein spots were matched to Coomassie stained gels run in parallel, cut out and trypsin digested. Matrix-assisted laser desorption/ionization time of flight-mass spectrometry (MALDI-TOF-MS) was used to determine the peptide mass fingerprints, which were searched in the MASCOT database to identify the protein.
A total of 8 immunoreactive proteins were successfully identified from the Coomassie stained gels in three repeats. Six of these proteins have not previously been reported as immunogenic in S. flexneri. These proteins could be potential candidates for vaccine or attenuation studies.
Soluble and membrane proteins of S. flexneri 2457T have been screened by 2-DE and immunoblotting with sera from shigellosis patients. Eight proteins are identified as immunogenic.
采用蛋白质组学方法分析人类感染期间福氏志贺菌表达的免疫原性蛋白。
对福氏志贺菌2457T的可溶性蛋白和膜蛋白提取物进行二维凝胶电泳（2-DE）分离。将蛋白质转移至聚偏二氟乙烯（PVDF）膜上，并用志贺菌病患者的血清进行免疫印迹。将反应性蛋白点与平行运行的考马斯亮蓝染色凝胶进行匹配，切下并进行胰蛋白酶消化。使用基质辅助激光解吸/电离飞行时间质谱（MALDI-TOF-MS）确定肽质量指纹图谱，并在MASCOT数据库中进行搜索以鉴定蛋白质。
在三次重复实验中，从考马斯亮蓝染色凝胶中成功鉴定出总共8种免疫反应性蛋白。其中6种蛋白此前未被报道为福氏志贺菌的免疫原性蛋白。这些蛋白可能是疫苗或减毒研究的潜在候选物。
通过二维凝胶电泳和用志贺菌病患者血清进行免疫印迹，对福氏志贺菌2457T的可溶性蛋白和膜蛋白进行了筛选。鉴定出8种具有免疫原性的蛋白。
