The apparent deglycase activity of DJ-1 results from the conversion of free methylglyoxal present in fast equilibrium with hemithioacetals and hemiaminals

Abstract

Edited by Ruma Banerjee 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 ϳ500 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-1mediated 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.

DJ-1: A promising therapeutic candidate for ischemia-reperfusion injury

“…In this scenario, DJ-1 has been described to convert MGO and GO in less harmful species, by acting as a glutathione-independent glyoxalase [ [40] , [41] , [42] , [43] , [44] ]. Furthermore, some studies have reported that DJ-1 can deglycate both proteins and nucleic acids upon reaction with MGO [ [45] , [46] , [47] , [48] , [49] , [50] ], though this function has proved contentious by other reports [ 51 , 52 ]. Interestingly, DJ-1 also seems to participate in the preservation of glucose homeostasis, by regulating the glucose levels in the brown adipose tissue [ 53 ] and by modulating the subcellular localization of the glycolytic enzyme hexokinase I [ 32 ].…”

Impact of DJ-1 and Helix 8 on the Proteome and Degradome of Neuron-Like Cells

“…The authors suggested that deglycation constitutes the primary cell-physiological function of DJ-1. However, this concept has been challenged [ 42 , 43 ]. Using the in vitro model system of differentiated SH SY5Y cells, we probed for the potential deglycation properties of the four different DJ-1 variants.…”

“…A protective effect of DJ-1 against protein glycation has been shown in yeast, bacteria [ 49 ] and human keratinocytes [ 50 ]. Whether DJ-1 detoxifies free MGO by acting as a glyoxalase [ 12 , 42 , 51 ] or removes adducts formed by MGO on proteins by acting as a deglycase [ 14 , 52 ] is a matter of debate. Here we confirm a C106-dependent protective effect of DJ-1 on protein glycation in neuron-like cells and show that in our cellular system H8 is essential for this effect.…”

The Role of DJ-1 in Cellular Metabolism and Pathophysiological Implications for Parkinson’s Disease

“…Loss of DJ-1 was shown to increase the levels of glycated DNA and DNA strand breaks [38]. However, the proposed deglycase activity of DJ-1 is not yet commonly accepted, as conflicting observations have been reported [108,109]. If confirmed, it would imply, however, that lack of functional DJ-1 could lead to reduced protection from glycation, increased DNA and protein damage and, hence, premature cellular aging via accumulation of advanced glycation end products (AGEs) [38].…”