Functional switching of a novel prokaryotic 2-Cys peroxiredoxin (PpPrx) under oxidative stress

Abstract

Many proteins have been isolated from eukaryotes as redox-sensitive proteins, but whether these proteins are present in prokaryotes is not clear. Redox-sensitive proteins contain disulfide bonds, and their enzymatic activity is modulated by redox in vivo. In the present study, we used thiol affinity purification and mass spectrometry to isolate and identify 19 disulfide-bond-containing proteins in Pseudomonas putida exposed to potential oxidative damages. Among these proteins, we found that a typical 2-Cys Prx-like protein (designated PpPrx) displays diversity in structure and apparent molecular weight (MW) and can act as both a peroxidase and a molecular chaperone. We also identified a regulatory factor involved in this structural and functional switching. Exposure of pseudomonads to hydrogen peroxide (H(2)O(2)) caused the protein structures of PpPrx to convert from high MW complexes to low MW forms, triggering a chaperone-to-peroxidase functional switch. This structural switching was primarily guided by the thioredoxin system. Thus, the peroxidase efficiency of PpPrx is clearly associated with its ability to form distinct protein structures in response to stress.

Insights into the Function of a Second, Nonclassical Ahp Peroxidase, AhpA, in Oxidative Stress Resistance in Bacillus subtilis

“…While this chaperone activity has mostly been associated with eukaryotic peroxiredoxins, it was shown that AhpC (a nonclassical Ahp) from Helicobacter pylori acts as a molecular chaperone under conditions of oxidative stress (69). Peroxiredoxins from Pseudomonas putida and P. aeruginosa were also shown to have chaperone activity (70)(71)(72). Consistent with this hypothesis, it was recently found that B. subtilis AhpA is inactivated in vitro when exposed to moderately high levels of H 2 O 2 (293 M); AhpC was unaffected by this level of peroxide (29).…”

Engineering of 2-Cys Peroxiredoxin for Enhanced Stress-Tolerance

“…Based on the enzymatic analysis results, this PpPrx has dual functions as a dominant chaperone and a recessive peroxidase activity (An et al, 2011). The PaPrx protein from P. aeruginosa PAO1 also contains 200 amino acids, and has dual functions as a dominant peroxidase and a recessive chaperone (An et al, 2010).…”

“…Oxidized Cys P is attacked by the resolving Cys (Cys R ). Two oxidized active Cys residues are formed via intra-or intermolecular disulfide bonds after yielding water or the corresponding alcohol (An et al, 2010;2011;Dietz, 2003;Hofmann et al, 2002).…”

“…Prxs can act either as peroxidases or as molecular chaperones, with reversible switching between the dual enzymatic functions (An et al, 2010;2011;Chuang et al, 2006;Jang et al, 2004;2006a). These properties enable Prxs to prevent the stress-induced misfolding or aggregation of intracellular macromolecules (Chuang et al, 2006;Jang et al, 2004;2006a).…”

Myo‐inositol‐1‐phosphate synthase (Ino‐1) functions as a protection mechanism in Corynebacterium glutamicum under oxidative stress

“…The oligomerization state of Ino‐1 was analyzed using the method previously described (An et al., ). The mixtures of the purified Ino‐1 and the loading buffer [250 mM Tris‐HCl (pH 6.8), 0.5% bromophenol blue (BPB), and 50% (v/v) glycerol] were separated on 12% PAGE and stained with Coomassie Brilliant Blue or detected by western blotting with anti‐his antibody.…”