An Essential Antioxidant in H. thermophilus Figure 3. Growth curves of wild-type (WT) H. thermophilus and the fpx gene disrupt mutant (Dfpx). Growth profiles of WT and Dfpx were observed under aerobic (A), anaerobic (B), and microaerobic (C) conditions. The WT strain and Dfpx were cultivated in inorganic medium at 70uC with shaking under aerobic (10% O2), anaerobic (0% O2, denitrification), and microaerobic (2% O2) conditions for 20 h. Optical density (OD) at 540 nm was monitored at 2-h intervals. Each OD was calculated based on the measurements of triplicate samples. Error bars show the standard deviations of each triplicate values. Closed and open circles denote WT and Dfpx, respectively. doi:10.1371/journal.pone.0034825.g003 observation of survival profiles of Dfpx in the presence of ROS, it was clearly demonstrated that the tolerance against ROS significantly decreased by defect of Fpx-system, strongly suggesting that Fpx plays a central role in the oxidative stress defense system of H. thermophilus. Although H. thermophilus utilizes the RTCA cycle, which is highly sensitive to oxidative condition, this bacterium grows preferably under aerobic conditions. Thus, Fpx may enable H. thermophilus to adopt such an unusual aerobic growth strategy. To our knowledge, this represents the first report to clarify the function of a single- domain Rbr-like protein as an antioxidant, and to reveal the essentiality of an Rbr-like protein for survival in aerobic environ- ments. The Km value of Fpx for H2O2 was greater than 40-fold higher than those of Pyrococcus furiosus Rbr (35 mM) and Clostridium acetobutylicum Rpr (,1 mM), and was similar to that of Anabaena sp. Rbr (2.25 mM) [6,25,26]. The high Km value of Fpx may imply its Figure 4. Survivals of WT H. thermophilus and Dfpx in the presence of ROS. Survival profiles of WT (A, B) and Dfpx (C, D) were observed under anaerobic (0% O2, denitrification) conditions, in the presence of paraquat (A, C) or H2O2 (B, D). Error bars show the standard deviations of each duplicate value. For panels A and C, open circle, closed circle, open triangle, closed triangle, open square denote 0, 1, 5, 10, and 100 mM paraquat. For panels B and D, open circle, closed circle, open triangle, closed triangle, open square denote 0, 10, 50, 100, and 500 mM H2O2. doi:10.1371/journal.pone.0034825.g004 PLoS ONE | www.plosone.org 4 April 2012 | Volume 7 | Issue 4 | e34825