These data pointed to the disparate metabolic networks operative in these systems and to the possible accumulation of KG and its utilization in combating oxidative stress. Roscovitine manufacturer It has been shown that KG is involved in the detoxification

of ROS with the concomitant formation of succinate. Ketoacids are known to eliminate ROS in a nonenzymatic manner (Brookes et al., 2006; Fedotcheva et al., 2006). Hence, it is not unlikely that P. fluorescens reprogrammed its metabolism in an effort to generate KG during the challenge posed by H2O2. This ketoacid has been shown to contribute to a decrease in oxidative tension (Li et al., 2009). The increased presence of succinate and KG in stressed cells would point to such a possibility. As KG was an important metabolite during oxidative stress, its utilization and production were monitored. ICDH, KGDH, and GDH are the three main participants in modulating the concentration of KG. In this study, there was a sharp increase in ICDH-NADP with a concomitant decrease in KGDH in the cells challenged by H2O2. As histidine was the only source of nitrogen and a possible precursor of KG, the presence of GDH-NAD and GDH-NADP was investigated. Although GDH-NADP was barely discernable in the control cells, there was a marked increase

in the H2O2-stressed cells. While LDK378 there was a mild increase in GDH-NAD, ICDH-NAD was sharply decreased in the H2O2-challenged cells. This is not surprising as NADH, a pro-oxidant, is known to further exacerbate the oxidative burden of the cell (Finkel & Holbrook, 2000; Thomas et al., 2009). Hence, the H2O2-stressed P. fluorescens may

have downregulated its formation. However, the upregulation of the NADPH production will be beneficial as this moiety plays a pivotal role in maintaining the reductive force of the microorganism during oxidative stress. Furthermore, the enhancement of these to enzymatic reactions (ICDH-NADP and GDH-NADP) will lead to the production of KG (Mailloux et al., 2009a, b). The decrease of KGDH has the net effect of increasing the pool of KG, a key contributor to the elimination of H2O2 (Brookes et al., 2006; Fedotcheva et al., 2006). Furthermore, the KGDH-mediated reaction has been shown to generate ROS (Starkov et al., 2004). To ascertain that the direct interaction between histidine and H2O2 does not lead to KG production, the growth medium with added H2O2 was monitored for 48 h without P. fluorescens. No KG was discerned (data not included). Hence, its downregulation will quell the oxidative burden of the microorganism, and limit the synthesis of NADH, a pro-oxidant. Thus, the enhanced activities of ICDH-NADP and GDH-NADP, coupled with the decreased activity of ICDH-NAD and KGDH, help generate KG and NADPH, two key ingredients necessary for survival during oxidative stress. As glutamate was an important supplier of KG, it was important to evaluate the status of other enzymes involved in the utilization or the formation of this substrate.