Abstract
Peroxiredoxins (Prdxs) are antioxidant enzymes that catalyse the breakdown of peroxides and regulate redox activity in the cell. Peroxiredoxin 5 (Prdx5) is a unique member of Prdxs, which displays a wider subcellular distribution and substrate specificity and exhibits a different catalytic mechanism when compared to other members of the family. Here, the role of a key metabolic integrator coenzyme A (CoA) in modulating the activity of Prdx5 was investigated. We report for the first time a novel mode of Prdx5 regulation mediated via covalent and reversible attachment of CoA (CoAlation) in cellular response to oxidative and metabolic stress. The site of CoAlation in endogenous Prdx5 was mapped by mass spectrometry to peroxidatic cysteine 48. By employing an in vitro CoAlation assay, we showed that Prdx5 peroxidase activity is inhibited by covalent interaction with CoA in a dithiothreitol-sensitive manner. Collectively, these results reveal that human Prdx5 is a substrate for CoAlation in vitro and in vivo, and provide new insight into metabolic control of redox status in mammalian cells.
Highlights
Peroxiredoxins (Prdxs) are antioxidant enzymes that neutralise a broad range of reactive oxygen and nitrogen species, including H2O2, alkyl hydroperoxides and peroxynitrites
We found that H2O2 perfusion significantly increased protein CoAlation when compared to control, and this was reversed to the level of untreated control by DTT (Fig. 1b)
We report for the first-time covalent modification of Peroxiredoxin 5 (Prdx5) by Coenzyme A (CoA) in cellular response to oxidative and metabolic stress
Summary
Peroxiredoxins (Prdxs) are antioxidant enzymes that neutralise a broad range of reactive oxygen and nitrogen species, including H2O2, alkyl hydroperoxides and peroxynitrites. Prdxs exhibit different subcellular localisation, regulation and substrate specificity. All members of the Prdx family share a common mechanism of peroxide-reducing activity that involves oxidation of a conserved peroxidatic cysteine (CysP) to a sulphenic acid (R-SOH), but differ in the mode. The activity and cellular functions of Prdxs are tightly controlled by regulatory interactions and post-translational modifications. A diverse range of binding partners for Prdxs have been identified, including proteins involved in signal transduction, metabolic processes and antioxidant defence [3,4,5]. Reversible thiol modifications, including S-sulphenylation, S-sulphinylation, S-glutathionylation and S-nitrosylation, as well as phosphorylation and acetylation were shown to play key roles in regulating peroxidase activity, subcellular localisation and redox signalling of Prdx family members [3,4,5,6]. By regulating intracellular peroxide levels, Prdxs were shown to participate in the regulation of cell proliferation, differentiation and redox signalling under physiological and pathological conditions [1, 7, 8]
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