Wang Ge, Maier Robert J
Department of Microbiology, University of Georgia, Athens, Georgia, USA.
Department of Microbiology, University of Georgia, Athens, Georgia, USA
J Bacteriol. 2015 Mar;197(5):973-82. doi: 10.1128/JB.02489-14. Epub 2014 Dec 22.
The gastric pathogen Helicobacter pylori must combat chronic acid and oxidative stress. It does so via many mechanisms, including macromolecule repair and gene regulation. Mitomycin C-sensitive clones from a transposon mutagenesis library were screened. One sensitive strain contained the insertion element at the locus of hp119, a hypothetical gene. No homologous gene exists in any (non-H. pylori) organism. Nevertheless, the predicted protein has some features characteristic of histone-like proteins, and we showed that purified HP119 protein is a DNA-binding protein. A Δhp119 strain was markedly more sensitive (viability loss) to acid or to air exposure, and these phenotypes were restored to wild-type (WT) attributes upon complementation of the mutant with the wild-type version of hp119 at a separate chromosomal locus. The mutant strain was approximately 10-fold more sensitive to macrophage-mediated killing than the parent or the complemented strain. Of 12 mice inoculated with the wild type, all contained H. pylori, whereas 5 of 12 mice contained the mutant strain; the mean colonization numbers were 158-fold less for the mutant strain. A proteomic (two-dimensional PAGE with mass spectrometric analysis) comparison between the Δhp119 mutant and the WT strain under oxidative stress conditions revealed a number of important antioxidant protein differences; SodB, Tpx, TrxR, and NapA, as well as the peptidoglycan deacetylase PgdA, were significantly less expressed in the Δhp119 mutant than in the WT strain. This study identified HP119 as a putative histone-like DNA-binding protein and showed that it plays an important role in Helicobacter pylori stress tolerance and survival in the host.
胃部病原体幽门螺杆菌必须应对慢性酸和氧化应激。它通过多种机制来做到这一点，包括大分子修复和基因调控。对转座子诱变文库中的丝裂霉素C敏感克隆进行了筛选。其中一个敏感菌株在假定基因hp119的位点含有插入元件。在任何（非幽门螺杆菌）生物体中都不存在同源基因。然而，预测的蛋白质具有一些组蛋白样蛋白质的特征，并且我们表明纯化的HP119蛋白是一种DNA结合蛋白。Δhp119菌株对酸或空气暴露明显更敏感（活力丧失），并且在突变体在单独的染色体位点用野生型hp119进行互补后，这些表型恢复到野生型（WT）属性。与亲本或互补菌株相比，突变菌株对巨噬细胞介导的杀伤的敏感性大约高10倍。在接种野生型的12只小鼠中，所有小鼠都含有幽门螺杆菌，而在12只小鼠中有5只含有突变菌株；突变菌株的平均定殖数量少158倍。在氧化应激条件下对Δhp119突变体和WT菌株进行蛋白质组学（二维PAGE与质谱分析）比较，发现了许多重要的抗氧化蛋白差异；SodB、Tpx、TrxR和NapA，以及肽聚糖脱乙酰酶PgdA，在Δhp119突变体中的表达明显低于WT菌株。这项研究将HP119鉴定为一种假定的组蛋白样DNA结合蛋白，并表明它在幽门螺杆菌的应激耐受性和在宿主体内的存活中起重要作用。
