Institute of Molecular Pathology, Faculty of Military Health Sciences, University of Defence, 500 01 Hradec Kralove, Czech Republic.
Department of Biochemical Studies, Faculty of Pharmacy in Hradec Kralove, Charles University in Prague, 500 05 Hradec Kralove, Czech Republic.
Microbiology (Reading). 2013 Nov;159(Pt 11):2364-2374. doi: 10.1099/mic.0.070516-0. Epub 2013 Sep 6.
Francisella tularensis is a highly infectious facultative intracellular bacterium and aetiological agent of tularaemia. The conserved hypothetical lipoprotein with homology to thiol/disulphide oxidoreductase proteins (FtDsbA) is an essential virulence factor in F. tularensis. Its protein sequence has two different domains: the DsbA_Com1_like domain (DSBA), with the highly conserved catalytically active site CXXC and cis-proline residue; and the domain amino-terminal to FKBP-type peptidyl-prolyl isomerases (FKBP_N). To establish the role of both domains in tularaemia infection models, site-directed and deletion mutagenesis affecting the active site (AXXA), the cis-proline (P286T) and the FKBP_N domain (ΔFKBP_N) were performed. The generated mutations led to high attenuation with the ability to induce full or partial host protective immunity. Recombinant protein analysis revealed that the active site CXXC as well as the cis-proline residue and the FKBP_N domain are necessary for correct thiol/disulphide oxidoreductase activity. By contrast, only the DSBA domain (and not the FKBP_N domain) seems to be responsible for the in vitro chaperone activity of the FtDsbA protein.
土拉弗朗西斯菌是一种高度传染性的兼性细胞内细菌，也是土拉菌病的病原体。与硫醇/二硫键氧化还原酶蛋白同源的保守假脂蛋白（FtDsbA）是土拉弗朗西斯菌的一个重要毒力因子。其蛋白序列有两个不同的结构域：DsbA_Com1 样结构域（DSBA），具有高度保守的催化活性位点 CXXC 和顺式脯氨酸残基；以及 FKBP 型肽脯氨酰基顺反异构酶的氨基端结构域（FKBP_N）。为了确定这两个结构域在土拉菌病感染模型中的作用，进行了影响活性位点（AXXA）、顺式脯氨酸（P286T）和 FKBP_N 结构域（ΔFKBP_N）的定点和缺失突变。产生的突变导致高衰减，具有诱导完全或部分宿主保护性免疫的能力。重组蛋白分析表明，活性位点 CXXC 以及顺式脯氨酸残基和 FKBP_N 结构域对于正确的硫醇/二硫键氧化还原酶活性是必需的。相比之下，只有 DSBA 结构域（而不是 FKBP_N 结构域）似乎负责 FtDsbA 蛋白的体外伴侣活性。
