Non-Mammalian Prdx6 Enzymes (Proteins with 1-Cys Prdx Mechanism) Display PLA₂ Activity Similar to the Human Orthologue.

Renata Bannitz-Fernandes,Rogério Aleixo-Silva,Chandra Dodia,Jose Vazquez-Medina,João Silva,Aron Fisher,Jian-Qin Tao,Luis Netto

doi:10.3390/antiox8030052

Renata Bannitz-Fernandes, Rogério Aleixo-Silva + Show 6 more

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https://doi.org/10.3390/antiox8030052

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Abstract

Mammalian peroxiredoxin class 6 (Prdx6) are bifunctional enzymes. Non-mammalian Prdx6 enzymes display Cys-based peroxidase activity, but to date their putative phospholipase A2 (PLA2 activities) has not been experimentally investigated. Initially, we observed that five non-mammalian Prdx6 enzymes (enzymes from Arabidopsis thaliana (AtPER1), Triticum aestivum (TaPER1), Pseudomonas aeruginosa (PaLsfA) and Aspergillus fumigatus (AfPrx1 and AfPrxC)) present features compatible with PLA2 activities in mammalian Prdx6 by amino acid sequences alignment and tertiary structure modeling. Employing unilamellar liposomes with tracer amounts of [3H]-1,2-Dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) and thin layer chromatography, all the tested non-mammalian Prdx6 enzymes displayed PLA2 activities, with values ranging from 3.4 to 6.1 nmol/min/mg protein. It was previously shown that Thr177 phosphorylation of human Prdx6 increases its PLA2 activity, especially at neutral pH. Therefore, we investigated if human Erk2 kinase could also phosphorylate homologous Thr residues in non-mammalian Prdx6 proteins. We observed phosphorylation of the conserved Thr in three out of the five non-mammalian Prdx enzymes by mass spectrometry. In the case of the mitochondrial Prdx6 from A. fumigatus (AfPrxC), we also observed phosphorylation by western blot, and as a consequence, the PLA2 activity was increased in acidic and neutral conditions by the human Erk2 kinase treatment. The possible physiological meanings of these PLA2 activities described open new fields for future research.

Highlights

Peroxiredoxins (Prdxs) are ubiquitous enzymes that play prominent roles in regulating peroxide levels within cells
Comparing the sequences of peroxiredoxin class 6 (Prdx6) from different organisms, we observed that the peroxidase motif (PVCTTE) (Figure 1, light red) and the corresponding catalytic triad (Figure 1, dark red) are fully conserved, with the exception of one single replacement of amino acid with similar physico-chemical properties in Prdx6 from Selaginella moellendorffii (PVCTSE)
It was realized that these a Ca2+ -independent phospholipase A2 (aiPLA2) and the human Prdx6 peroxidase activity of human Prdx6 was described

Summary

Introduction

Peroxiredoxins (Prdxs) are ubiquitous enzymes that play prominent roles in regulating peroxide levels within cells. These enzymes are often abundant and are capable of rapidly reducing peroxides [1]. Prdxs are Cys-based peroxidases that contain one or two conserved Cys residues [2]. All characterized Prdxs display the universal motif PxxxT/SxxC and can reduce hydroperoxides efficiently, with rates ranging from 106 to 108 M−1 s−1 [3]. In spite of being a heterogeneous family, all Prdxs have this peroxidase activity dependent on Thr(Ser)-Cys-Arg catalytic triad that stabilizes the transition state of the reaction [4].

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