Optimized enzymatic dual functions of PaPrx protein by proton irradiation

Abstract

We investigated the effects of proton irradiation on the function and structure of the Pseudomonas aeruginosa peroxiredoxin (PaPrx). Polyacrylamide gel demonstrated that PaPrx proteins exposed to proton irradiation at several doses exhibited simultaneous formation of high molecular weight (HMW) complexes and fragmentation. Size-exclusion chromatography (SEC) analysis revealed that the number of fragments and very low molecular weight (LMW) structures increased as the proton irradiation dose increased. The peroxidase activity of irradiated PaPrx was preserved, and its chaperone activity was significantly increased by increasing the proton irradiation dose. The chaperone activity increased about 3–4 fold after 2.5 kGy proton irradiation, compared with that of non-irradiated PaPrx, and increased to almost the maximum activity after 10 kGy proton irradiation. We previously obtained functional switching in PaPrx proteins, by using gamma rays and electron beams as radiation sources, and found that the proteins exhibited increased chaperone activity but decreased peroxidase activity. Interestingly, in this study we newly found that proton irradiation could enhance both peroxidase and chaperone activities. Therefore, we can suggest proton irradiation as a novel protocol for conserved 2-Cys protein engineering.

Site-directed mutagenesis substituting cysteine for serine in 2-Cys peroxiredoxin (2-Cys Prx A) ofArabidopsis thalianaeffectively improves its peroxidase and chaperone functions

“…However, earlier attempts to manipulate 2-Cys Prx to enhance one of these activities have compromised the efficiency of the other activity (An et al, 2011a;Hong et al, 2012). Recently, however, Park et al (2014) have reported that proton irradiation can enhance both the peroxidase and the chaperone activity of PaPrx (2-Cys Prx). It has also been observed that the dual function of 2-Cys Prx can be regulated by an additional Cys on the a-helix region between two active Cys residues (An et al, 2011b).…”