The NreA/NreB sensors of Staphylococcus carnosus: A sensor complex for coordinated-sensing of O2 and nitrate.

细菌对环境刺激反应迅速，并调整细胞组成、代谢和其他特性以达到最佳生长和生存。电子受体如O2和硝酸盐是控制许多基因表达的许多细菌的主要刺激物，包括有氧和无氧呼吸，发酵，以及毒力和生物技术相关性状。在葡萄球菌对O2和硝酸盐的反应中，出现了使用新成分和遵循新策略的情况。氧传感器NreB是细胞质双组分系统（NreB/NreC）的一部分。NreB传感器激酶由一个感觉PAS结构域组成，该结构域结合一个不稳定的[4Fe-4S]2+簇进行O2传感，该簇被O2降解为[2Fe-2S]2+簇，然后完全丢失，形成apoNreB。后一种形式在硝酸盐呼吸基因的转录激活中不活跃。硝酸盐受体NreA是一个小的细胞质GAF结构域蛋白，在活性位点结合硝酸盐。与T. Stehle （t<s:1> bingen）合作对该新型硝酸盐受体的结构进行了表征。在美因茨，O2和硝酸盐传感之间的相互作用仍在继续。NreA与NreB直接相互作用，相互作用受硝酸盐控制。无硝酸盐的NreA与NreB结合，然后在自磷酸化中被抑制。因此，NreC的磷酸化和靶基因的表达减少。当NreA以硝酸盐结合形式存在时，NreA/NreB的相互作用和对NreB磷酸化的抑制作用得以缓解。因此，NreA和NreB在NreA/NreB传感器复合物中合作，其中主传感器NreB的活性由硝酸盐受体NreA调节；NreA不能直接控制靶基因。这种O2/硝酸盐感知模式与大肠杆菌、芽孢杆菌、假单胞菌等其他细菌的感知模式有着根本的区别。新型NreA/NreB传感器复合物将在分子水平上表征其在NreA/NreB相互作用以及NreA和硝酸盐对o2传感器NreB的控制方面的特性。因此，(i) NreA和NreB上的物理相互作用位点，以及（ii） NreB中受NreA调节的分子内信号传递位点（FeS簇，PAS结构域，激酶结构域）应在分子水平上确定。此外（iii）， NreB含有一个活跃的磷酸酶结构域，这将以其对NreB活性的作用为特征。NreB的结构研究（iv）已经开始并将继续进行（与Homburg的R. Lancaster合作）。

Bacteria respond rapidly to environmental stimuli, and adapt cellular composition, metabolism and other properties to optimal growth and survival. Electron acceptors like O2 and nitrate represent major stimuli for many bacteria that control expression of many genes, including those for aerobic and anaerobic respiration, fermentation, but also virulence and biotechnologically relevant traits. In Staphylococci response to O2 and nitrate turned out to use new components and to follow new strategies. The oxygen sensor NreB is part of a cytoplasmic two-component system (NreB/NreC). The NreB sensor kinase consists of a sensory PAS domain that binds a labile [4Fe-4S]2+ cluster for O2-sensing which is degraded by O2 to a [2Fe-2S]2+ cluster, and then completely lost, forming apoNreB. The latter forms are inactive in transcriptional activation of the genes of nitrate respiration. The nitrate receptor NreA is a small cytoplasmic GAF domain protein that binds nitrate at the active site. The structure of this novel nitrate receptor was characterized in collaboration with T. Stehle (Tübingen). Interaction between O2 and nitrate sensing was continued in Mainz. NreA interacts directly with NreB, and the interaction is controlled by nitrate. Nitrate-free NreA binds to NreB which is then inhibited in autophosphorylation. Consequently, phosphorylation of NreC and expression of target genes is decreased. The NreA/NreB interaction and inhibition of NreB phosphorylation is relieved when NreA exists in the nitrate-bound form. Thus NreA and NreB cooperate in a NreA/NreB sensor complex, where the primary sensor NreB is modulated in activity by the nitrate-receptor NreA; NreA is not able of directly controlling target genes. This mode of O2/nitrate sensing is in fundamental contrast to that of other bacteria like E. coli, Bacillus, Pseudomonas.The novel NreA/NreB sensor complex will be characterized at the molecular level for its characteristic properties regarding NreA/NreB interaction and control of the O2-sensor NreB by NreA and nitrate. Thus (i) the physical interaction sites on NreA and NreB, and (ii) the sites in NreB for the intramolecular signal transfer (FeS cluster, PAS domain regions, kinase domain) that are modulated by NreA, shall be identified at the molecular level. In addition (iii), NreB contains an active phosphatase domain, which will be characterized in its role for NreB activity. Structural studies (iv) on NreB have been started and will be continued (collaboration with R. Lancaster, Homburg).