Oxidizable Residues Mediating Protein Stability and Cytoprotective Interaction of DJ-1 with Apoptosis Signal-regulating Kinase 1

Abstract

2 (1). The most dramatic PD-associated mutation L166P impairs DJ-1 dimer formation and dramatically destabilizes the protein (2-7). Other mutations such as M26I (8) and E64D (9) have more subtle defects with unclear cellular consequences (4, 7, 10, 11). In addition to this genetic association, DJ-1 is neuropathologically linked to PD. DJ-1 is up-regulated in reactive astrocytes, and it is oxidatively modified in brains of sporadic PD patients (12)(13)(14).DJ-1 protects against oxidative stress and mitochondrial toxins in cell culture (15-17) as well as in diverse animal models (18 -21). The cytoprotective effects of DJ-1 may be stimulated by oxidation and mediated by molecular chaperoning (22, 23), and/or facilitation of the pro-survival Akt and suppression of apoptosis signal-regulating kinase 1 (ASK1) pathways (6,24,25). The cytoprotective activity of DJ-1 against oxidative stress depends on its cysteine residues (15,17,26). Among the three cysteine residues of DJ-1, the most prominent one is the easiest oxidizable that is in a constrained conformation (28), but the other cysteine residues Cys-46 and Cys-53 have been implicated with DJ-1 activity as well (22). However, the molecular basis of oxidation-mediated cytoprotective activity of DJ-1 is not clear. Moreover, the roles of PD-mutated and in vivo oxidized methionines are not known.Here we have mutagenized all oxidizable residues within DJ-1 and studied the effects on protein stability and function. The PD-associated mutation M26I within the DJ-1 dimer interface selectively reduced protein expression as well as ASK1 suppression and cytoprotective activity in oxidatively stressed cells. These cell culture results support a pathogenic effect of the clinical M26I mutation (8). Furthermore, oxidation-defective C106A mutation abolished binding to ASK1 and cytoprotective activity of DJ-1, whereas the designed higher order oxidation

Dopamine-derived Quinones Affect the Structure of the Redox Sensor DJ-1 through Modifications at Cys-106 and Cys-53

“…Mutant proteins DJ-1(C53A) and DJ-1(C106A) were cloned and purified. The single mutation of Cys-46 (C46A) produces an unstable protein, which is prone to degradation (25,34), and the C46S, which we cloned and purified, is more unstable than the other single mutant proteins. Therefore, to obtain information on Cys-46 indirectly, a double mutant, DJ-1(C53A/C106A), was prepared.…”

“…Therefore, the full cytoprotective activity of DJ-1 seems to require a more complete, mixed disulfide-mediated incorporation into the ASK1 signalosome, for which Cys-106 is necessary. It has been suggested that the C106DD mutation only "opens" the conformation of DJ-1 to reveal ASK1 binding site(s), resulting in the observed ASK1 binding and partial cytoprotection (34). Also this functional hypothesis requires Cys-106, which would be severely compromised by DAQ binding.…”

Park7 interacts with p47phox to direct NADPH oxidase-dependent ROS production and protect against sepsis

“…Remarkably, this interaction was enhanced in NT after LPS challenge in a time-dependent manner ( Figure 7A). As Cys106 oxidation is important for Park7 interaction with other proteins [37], we then examined whether Park7 was oxidized in our experimental system. NT cells showed significantly increased Park7 oxidation after PMA treatment (Supplementary information, Figure S8), suggesting that the enhanced Park7-p47 phox interaction may be attributed to the increased Park7 oxidation level.…”

The Redox Biochemistry of Protein Sulfenylation and Sulfinylation

“…Considering the high propensity of Cys-106 to oxidize, it has been proposed that DJ-1 acts merely as a direct ROS scavenger. However, an elegant new study reported that the DJ-1 C106DD mutant is still able to protect cells against oxidative stress (70), excluding direct scavenger action by Cys oxidation.…”