Function of a new sensor kinase NreB from Staphylococcus carnosus using a [4Fe4S]2+ cluster for sensing oxygen

肉质葡萄球菌使用细胞质NreBC双组分系统来感知O2和硝酸盐呼吸的转录调节。NreB含有一种新型的感觉PAS(Per-Arnt-Sim)结构域，它结合了[4Fe-4S]2+簇。该簇合物通过与O2反应降解，生成[2Fe-2S]2+作为中间体，生成(无FeS)apoNreB作为最终产物。ApoNreB和[2Fe-2S]2+.NreB失去了激酶活性和基因激活能力。结果表明，在需氧细菌中apoNreB是NreB的失活形式，而[2Fe-2S]2+。NreB没有明显作用。总体而言，NreB中的簇降解直接控制NreB的激酶活性，而在O2传感器FNR中，相同的反应控制蛋白质的二聚化和DNA结合。我们将研究NreB的FES结合部位和信号传递到激活域的过程，以便在蛋白质水平上了解FNR的分子差异。含有NreBC的细菌还含有基本上由GAF结构域组成的NreA蛋白。NreA在NreBC的O2传感中发挥重要作用，并与NreB发生物理作用。我们推测，NreA是一种辅助蛋白，控制NreB在O2传感中的功能，NreBC与NreA共同形成一个三组分系统。我们将研究NreA和GAF结构域在体内和体外对NreBC功能的作用。

Staphylococcus carnosus uses the cytoplasmic NreBC two-component system for sensing O2 and transcriptional regulation of nitrate respiration. NreB contains a new type of a sensory PAS (Per-Arnt-Sim) domain that binds a [4Fe-4S]2+ cluster. The cluster is degraded by reaction with O2, producing [2Fe-2S]2+ as an intermediate and (FeS-less) apoNreB as the final product. ApoNreB and [2Fe-2S]2+.NreB have lost the kinase activity and the capacity for gene activation. It is shown that apoNreB is the inactive form of NreB in aerobically grown bacteria whereas [2Fe-2S]2+.NreB plays no significant role. Overall, cluster degradation in NreB directly controls kinase activity of NreB, whereas in the O2-sensor FNR the same reaction controls dimerisation and DNA-binding of the protein. The processes at the FeS binding site of NreB and in the signal transfer to the kinase domain will be studied to understand the molecular differences to FNR at the protein level. Bacteria containing NreBC harbour in addition the NreA protein that consists essentially of a GAF domain. NreA fulfils an important role in O2-sensing by NreBC and physically interacts with NreB. We suppose that NreA functions as an accessory protein that controls the function of NreB in O2 sensing, and that NreBC forms together with NreA a three-component system. We will study the role of NreA and the GAF domain for NreBC function in vivo and in vitro.