Therefore, we decided to study the expression of these genes in greater detail. a. Regulation of sodA and sodB There is CBL0137 datasheet plethora of information about the regulation of sodA and sodB in E. coli [80–85], but there is little knowledge about the regulation of these genes in S. Typhimurium [86]. In the present study, the microarray data showed that the anaerobic expression of sodA and sodB

in Δfur was > 9-fold higher and > 3-fold lower, respectively, than in the parent WT strain (Additional file 2: Table S2). SodA (MnSOD) and SodB (FeSOD) are the cytosolic superoxide dismutases of S. Typhimurium and they require the cofactors manganese and iron, respectively. These SODs are homodimers, and are fully functional when metalated with the appropriate metals (i.e., manganese for SodA and iron for SodB). However, a heterodimer consisting of SodA(Mn)/SodB(Fe) TH-302 selleck kinase inhibitor can still exhibit SOD activity, albeit at a reduced level compared to the homodimer [87]. Thus, in order to see an active hybrid SOD, both SodA and SodB must be

expressed. Data in Figure 3A demonstrated that, as in anaerobic E. coli, the WT strain (Lane 1) lacked the activity of both Mn- and Hybrid-SODs, but possessed an active FeSOD. However, Δfur (Figure 3A – Lane 2) was devoid of all three SOD-isozymes. The lack of FeSOD in Δfur was of no surprise, as previous studies in E. coli [83, 84] have established that Fur is indirectly required for the translation of sodB via its repression of the small RNA, ryhB, which works in conjunction with the RNA chaperon protein, Hfq [88, 89]. Indeed, a strain harboring deletions in both Fur and Hfq (ΔfurΔhfq) resulted in restoration of SodB activity (Figure 3A – Lane 4). Furthermore, the high degree of sequence identity in the promoter and the gene sequence of ryhB of E. coli with the two ryhB-like small RNAs, rfrA and rfr of S. Typhimurium [39], suggested that the regulation of sodB in S. Typhimurium is similar to that reported in E. coli [88, 89]. Interestingly, expression of the hybrid SOD appears up-regulated in Δhfq and ΔfurΔhfq clonidine (Figure 3A – Lane 3 and 4). The reason for this is unclear,

but may be due to the activation of the Hfq-binding small RNA (fnrS) by Fnr, which subsequently represses the expression of sodA [90, 91]. Figure 3 Effects of Fur, Hfq, and manganese on the activity of superoxide dismutases. (A) Effects of Fur and Hfq – Cell-free extracts from anaerobically grown cultures (14028s, Δfur, Δhfq, and ΔfurΔhfq) were prepared as described in the Methods. Equal protein (125 μg/ml) was loaded and following electrophoresis the gel was stained for SOD activity. Lane 1 – 14028s; lane 2 – Δfur; lane 3 – Δhfq; lane 4 – ΔfurΔhfq. (B) Effects of Fur and MnCl2 – Cell-free extracts were prepared from anaerobically grown cultures as in (A) except that 1 mM MnCl2 was added to the media. Equal protein (125 μg/lane) was loaded, elecrophoresed, and stained for SOD as in (A).