Center of Advanced Studies, Faculty of Military Health Science UO, 500 01 Hradec Kralove, Czech Republic.
J Proteome Res. 2009 Nov;8(11):5336-46. doi: 10.1021/pr900570b.
Francisella tularensis (F. tularensis) is highly infectious for humans via aerosol route and untreated infections with the highly virulent subsp. tularensis can be fatal. Our knowledge regarding key virulence determinants has increased recently but is still somewhat limited. Surface proteins are potential virulence factors and therapeutic targets, and in this study, we decided to target three genes encoding putative membrane lipoproteins in F. tularensis LVS. One of the genes encoded a protein with high homology to the protein family of disulfide oxidoreductases DsbA. The two other genes encoded proteins with homology to the VacJ, a virulence determinant of Shigella flexneri. The gene encoding the DsbA homologue was verified to be required for survival and replication in macrophages and importantly also for in vivo virulence in the mouse infection model for tularemia. Using a combination of classical and shotgun proteome analyses, we were able to identify several proteins that accumulated in fractions enriched for membrane-associated proteins in the dsbA mutant. These proteins are substrate candidates for the DsbA disulfide oxidoreductase as well as being responsible for the virulence attenuation of the dsbA mutant.
弗氏志贺样杆菌（F. tularensis）可通过气溶胶途径高度感染人类，未经治疗的高度毒力亚种感染可能致命。我们最近对关键毒力决定因素的了解有所增加，但仍有些有限。表面蛋白是潜在的毒力因子和治疗靶点，在这项研究中，我们决定针对弗氏志贺样杆菌 LVS 中编码三个假定膜脂蛋白的基因。其中一个基因编码的蛋白质与二硫键氧化还原酶 DsbA 家族的蛋白质具有高度同源性。另外两个基因编码的蛋白质与志贺氏菌的毒力决定因子 VacJ 具有同源性。编码 DsbA 同源物的基因被证明是在巨噬细胞中存活和复制所必需的，而且对于兔热病的小鼠感染模型中的体内毒力也很重要。使用经典和鸟枪法蛋白质组分析的组合，我们能够鉴定出在 dsbA 突变体中富含膜相关蛋白的部分中积累的几种蛋白质。这些蛋白质是 DsbA 二硫键氧化还原酶的底物候选物，也是 dsbA 突变体毒力衰减的原因。
