56 - Novel Redox-Dependent Esterase Activity for DJ-1: Implications for Parkinson's Disease

Abstract

Mutations in human DJ-1 (hDJ-1) are associated with early- onset autosomal recessive forms of Parkinson’s disease (PD). hDJ-1 is a multi-functional protein that functions as an oxidative sensor, transcription factor, protease, deglycase and glyoxalase. hDJ-1 contains a conserved cysteine protease domain. Given that cysteine proteases can act on both amide and ester substrates, we surmised that hDJ-1 possesses cysteine- mediated esterase activity. To test this hypothesis, hDJ-1 was overexpressed, purified and tested for esterase activity towards p- nitrophenyl-acetate (pNPA). hDJ-1 showed esterase activity (Vmax= 235.10 ± 12.00), with a sigmoidal fit (S 0.5 = 0.55 ±0.040 mM) and positive cooperativity (Hill-coefficient of 2.05 ± 0.28). A PD-associated mutant of DJ-1 (M26I) lacked activity. Unlike its protease activity which is inactivated by reactive oxygen species (ROS), esterase activity of hDJ-1 is enhanced upon exposure to low concentrations of hydrogen peroxide ( 100 µM) suggesting that its activity is resistant to oxidative stress. Esterase activity of DJ-1 requires the oxidation of catalytic cysteines, as chemically protecting cysteines blocks its activity whereas an oxido-mimetic mutant of DJ-1 (C106D) exhibited robust esterase activity. Redox-dependent activation of esterase activity is specific for DJ-1 as treating porcine pancreatic lipase, a serine protease, did not show enhanced esterase activity when treated with hydrogen peroxide. Finally, molecular docking studies suggested that C106 and L126 within the catalytic site of hDJ-1 interact with esterase substrates. Ongoing cell culture studies will elucidate the extent by which the esterase activity of DJ-1 plays a cytoprotective role in primary neurons treated with ROS (hydrogen peroxide). This study is the first to report intrinsic redox-sensitive esterase activity for hDJ-1. Overall, our results suggest that the esterase activity of DJ-1 plays a role in conferring cytoprotection during oxidative stress.