A transient post-translational modification of active site cysteine alters binding properties of the parkinsonism protein DJ-1

Mutations in the human protein DJ-1 cause early onset of Parkinson's disease. A reactive cysteine residue (Cys106) of DJ-1 is crucial for its protective function, although the underlying mechanisms are unclear. Here we show that a fraction of bacterially expressed polyhistidine-tagged human DJ-1 could not be eluted from a Ni-nitrilotriacetate (Ni-NTA) column with 150 mM imidazole. This unusually tight binding was accompanied by the appearance of blue violet color on the Ni-NTA column. We demonstrate by X-ray crystallography that Cys106 is carboxymethylated in a fraction of DJ-1 tightly bound to Ni-NTA and that the replacement of Cys106 by serine abrogates the tight binding and the appearance of blue violet color. However, carboxymethylation of purified DJ-1 is insufficient to confer the tight binding to Ni-NTA. Moreover, when eluted protein was re-applied to the Ni-NTA column, no tight binding was observed, indicating that the formation of high affinity complex with Ni-NTA depends on a transient modification of Cys106 that transforms into a Cys106-carboxymethyl adduct upon elution from Ni-NTA. We conclude that an unknown metabolite reacts with Cys106 of DJ-1 to result in a transient post-translational modification. This modification is distinct from simple oxidation to sulfinic or sulfenic acids and confers altered binding properties to DJ-1 suggesting that it could serve as a signal for sensing oxidant stress. •A fraction of bacterially expressed polyhistidine-tagged human DJ-1 tightly binds to Ni-nitrilotriacetate (Ni-NTA) column.•The color of Ni-NTA column changes from light cyan to blue violet upon tight binding to DJ-1.•An active site cysteine is post-translationally modified in the fraction of DJ-1 tightly bound to Ni-NTA.•The modificationconferring tightbinding to Ni-NTA is unstable.•Crystal structure shows the unstable modification converts to S-carboxymethylcysteine upon elution from Ni-NTA.