Denitrification and arginine fermentation are major parts of the anaerobic metabolism of Pseudomonas aeruginosa, which is important for biofilm formation and infection. The two-component regulatory system NarX-NarL is part of the underlying network and is required for denitrifying growth. All target promoters identified so far are activated by NarL. In this study the effect of NarL on arginine fermentation was investigated using proteome, Northern blot and lacZ reporter gene analyses. NarL-dependent repression of the arcDABC operon was observed and the corresponding NarL-binding site in the arcD promoter region was functionally localized at -60 bp upstream of the transcriptional start site using site-directed promoter mutagenesis and reporter gene fusion experiments. The results clearly show that in the presence of nitrate NarL represses the arginine-dependent activation of the arcDABC operon mediated by ArgR. It does not influence the oxygen-tension-dependent activation via Anr. Thus, the anaerobic energy metabolism of P. aeruginosa is coordinated via NarX-NarL activity. In the presence of nitrate the highly efficient denitrification is preferred over the less attractive arginine fermentation.

译文

反硝化和精氨酸发酵是铜绿假单胞菌厌氧代谢的主要组成部分,对生物膜的形成和感染具有重要意义。两部分监管系统narx-narl是基础网络的一部分,是反硝化增长所必需的。到目前为止,所有鉴定的靶启动子都被NarL激活。在这项研究中,使用蛋白质组,Northern印迹和lacZ报告基因分析研究了NarL对精氨酸发酵的影响。使用定点启动子诱变和报告基因融合实验,观察到arcDABC操纵子的NarL依赖性抑制,并且arcD启动子区域中相应的NarL结合位点功能定位在转录起始位点上游-60 bp。结果清楚地表明,在存在硝酸盐的情况下,NarL抑制arcDABC操纵子的精氨酸依赖性激活。它不会通过Anr影响氧张力依赖性激活。因此,铜绿假单胞菌的厌氧能量代谢是通过NarX-NarL活性协调的。在硝酸盐存在下,与吸引力较小的精氨酸发酵相比,高效的反硝化作用是优选的。

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