The up-regulation of tryptophan synthase in Pseudomonas sp. TLC6-6.5-4 in the presence of copper was consistent with our transposon mutational analysis (CSM1 trpA) (Table 1). The overexpression of trpA gene induced by copper treatment was reported in Helicobacter pylori (Waidner et al., 2002). Besides tryptophan,

our metabolomic analysis showed that the levels of several other amino acids such as l-proline and l-isoleucine were significantly increased when Pseudomonas sp. TLC6-6.5-4 was grown in the presence of 4 mM copper (Fig. 4), which correlates with the up-regulation of ketol-acid reductoisomerase, an enzyme involved in the biosynthesis of leucine and isoleucine (Table 1). An increase in amino acid synthesis was also identified in the multiple metal-resistant click here bacteria P. fluorescens in both biofilm and planktonic Selleckchem PD-332991 culture, which could be a protective

mechanism against enzyme inhibition or replacement of damaged proteins caused by the presence of copper (Booth et al., 2011). Furthermore, the accumulation of l-proline itself is the protective mechanism that bacteria (and plants and yeast) use to cope with the oxidative stress caused by heavy metals (Nandakumar et al., 2011). The Clp proteases play an important role in regulating cellular functions by refolding or degrading damaged proteins and also regulate the expression of genes involved in oxidative stress and DNA repair (Hengge & Bukau, 2003; Michel et al., 2006). However, very little is known about the role of Clp proteases in Pseudomonas species except for the basic function of proteolysis. Disruption of ClpA in P. putida CA-3 decreased polyhydroxyalkanoates, the intracellular granules,

in response to inorganic nutrient limitation (Goff et al., 2009). In the present study, we demonstrated that the transposon insertion mutant, CSM2, disrupted in Clp protease subunit ClpA showed a significant reduction in copper resistance compared with the wild-type strain. A recent study on Staphylococcus aureus also showed that the expression Etofibrate of ClpA was up-regulated in response to copper (Baker et al., 2010). The disruption of ClpA caused the down-regulation of glycosyl transferase and tRNA (guanine-N(7)-)-methyltransferase (Table 1). Glycosyl transferase is essential for bacterial biofilm formation and resistance to oxidative stress (Erb et al., 2009; Tao et al., 2010). The higher levels of tRNA methyltransferase under cellular stress response are likely to reduce the degradation of tRNAs by ribonucleases activated under stress conditions (Thompson & Parker, 2009; Chan et al., 2010). DnaJ-class molecular chaperone (Table 1), whose expression was up-regulated in wild-type strain grown with copper compared with wild type without copper, binds unfolded polypeptide chains, preventing their irreversible aggregation (Düppre et al.