Abstract
Loss-of-function mutations in the gene encoding human protein DJ-1 cause early onset of Parkinson's disease, suggesting that DJ-1 protects dopaminergic neurons. The molecular mechanisms underlying this neuroprotection are unclear; however, DJ-1 has been suggested to be a GSH-independent glyoxalase that detoxifies methylglyoxal (MGO) by converting it into lactate. It has also been suggested that DJ-1 serves as a deglycase that catalyzes hydrolysis of hemithioacetals and hemiaminals formed by reactions of MGO with the thiol and amino groups of proteins. In this report, we demonstrate that the equilibrium constant of reaction of MGO with thiols is ∼500 m-1 at 37 °C and that the half-life of the resulting hemithioacetal is only 12 s. These thermodynamic parameters would dictate that a significant fraction of free MGO will be present in a fast equilibrium with hemithioacetals in solution. We found that removal of free MGO by DJ-1's glyoxalase activity forces immediate spontaneous decomposition of hemithioacetals due to the shift in equilibrium position. This spontaneous decomposition of hemithioacetals could be mistaken for deglycase activity of DJ-1. Furthermore, we demonstrate that higher initial concentrations of hemithioacetals are associated with lower rates of DJ-1-mediated conversion of MGO, ruling out the possibility that hemithioacetals are DJ-1 substrates. Experiments with CRISPR/Cas-generated DJ-1-knockout HEK293 cells revealed that DJ-1 does not protect against acute MGO toxicity or carboxymethylation of lysine residues in cells. Combined, our results suggest that DJ-1 does not possess protein deglycase activity.
Highlights
Loss-of-function mutations in the gene encoding human protein DJ-1 cause early onset of Parkinson’s disease, suggesting that DJ-1 protects dopaminergic neurons
When DJ-1 was used instead of Glx3, no lactate was detected under the same conditions; when incubation time was increased to 10 h, lactate was readily detectable (Fig. 3A), demonstrating that DJ-1 exhibits a weak glyoxalase activity
Given the lack of consensus whether DJ-1 exerts its protective effects through an enzymatic detoxification of MGO and/or adducts of MGO with amino acids, we re-examined the glyoxalase and deglycase activities of DJ-1
Summary
Anna Andreeva‡, Zhanibek Bekkhozhin§, Nuriza Omertassova‡, Timur Baizhumanov§, Gaziza Yeltay‡, Mels Akhmetali‡, Daulet Toibazar‡, and X Darkhan Utepbergenov§1 From the Departments of ‡Biology and §Chemistry, School of Sciences and Humanities, Nazarbayev University, Nur-Sultan 010000, Kazakhstan
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