Department of Medicine, Centre for Molecular Medicine and Infection, Imperial College London, London, UK.
FEMS Microbiol Lett. 2012 May;330(1):38-45. doi: 10.1111/j.1574-6968.2012.02530.x. Epub 2012 Mar 12.
Nitrogen is an essential element required for bacterial growth and consequently bacteria must adapt to situations of nitrogen limitation for survival. The transcriptional response to nitrogen limitation in Mycobacterium smegmatis is thought to be regulated by GlnR, although, to date, only five nitrogen metabolism genes have been shown to be under its direct control. GlnR belongs to the OmpR family of two-component response regulators that are typically activated by phosphorylation of a conserved aspartate residue. The M. smegmatis GlnR protein contains the highly conserved aspartate residue (D48) corresponding to the phosphorylation sites identified in other OmpR family regulators. In this study, we replaced GlnR D48 with alanine and constructed a GlnR deletion mutant. Under nitrogen-limiting conditions, both the GlnR_D48A and GlnR deletion mutants exhibited reduced growth rates compared with wild type. Transcriptional analysis showed both mutants failed to up-regulate the expression of GlnR-controlled genes under nitrogen-limiting conditions. We therefore demonstrate that the GlnR aspartate (D48) residue is essential for its function as a nitrogen-stress transcriptional response regulator in M. smegmatis.
氮是细菌生长所必需的元素，因此细菌必须适应氮限制的生存环境。人们认为，Mycobacterium smegmatis 中对氮限制的转录反应受 GlnR 调控，尽管迄今为止，只有 5 个氮代谢基因被证明受其直接控制。GlnR 属于双组分反应调节剂的 OmpR 家族，通常通过磷酸化保守的天冬氨酸残基而被激活。M. smegmatis GlnR 蛋白含有高度保守的天冬氨酸残基 (D48)，与其他 OmpR 家族调节剂中鉴定的磷酸化位点相对应。在这项研究中，我们用丙氨酸取代了 GlnR D48，并构建了 GlnR 缺失突变体。在氮限制条件下，与野生型相比，GlnR_D48A 和 GlnR 缺失突变体的生长速度都有所降低。转录分析表明，这两种突变体在氮限制条件下都未能上调 GlnR 控制的基因的表达。因此，我们证明了 GlnR 中的天冬氨酸 (D48) 残基对于其作为 M. smegmatis 中氮胁迫转录反应调节剂的功能是必不可少的。
