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Kinetic evidence in favor of glyoxalase III and against deglycase activity of DJ-1.

ABSTRACT: DJ-1, a protein encoded by PARK7 plays a protective role against neurodegeneration. Since its glyoxalase III activity catalyzing methylglyoxal (MG) to lactate was discovered, DJ-1 has been re-established as a deglycase decomposing the MG-intermediates with amino acids and nucleotides (hemithioacetal and hemiaminal) rather than MG itself, but it is still debatable. Here, we have clarified that human DJ-1 directly recognizes MG, and not MG-intermediates, by monitoring the detailed catalytic processes and enantiomeric lactate products. The hemithioacetal intermediate between C106 of ¹⁵ N-labeled DJ-1 (^15N DJ-1) and MG was also monitored by NMR. TRIS molecule formed stable diastereotopic complexes with MG (K_d , 1.57 ± 0.27 mM) by utilizing its three OH groups, which likely disturbed the assay of deglycase activity. The low k_cat of DJ-1 for MG and its MG-induced structural perturbation may suggest that DJ-1 has a regulatory function as an in vivo sensor of reactive carbonyl stress.

SUBMITTER: Choi J

PROVIDER: S-EPMC10127264 | biostudies-literature | 2023 May

REPOSITORIES: biostudies-literature

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Kinetic evidence in favor of glyoxalase III and against deglycase activity of DJ-1.

Choi Joonhyeok J Tak Sungho S Jung Hoe-Myung HM Cha Soyoung S Hwang Eunha E Lee Donghan D Lee Joon-Hwa JH Ryu Kyoung-Seok KS Park Chankyu C

Protein science : a publication of the Protein Society 20230501 5

DJ-1, a protein encoded by PARK7 plays a protective role against neurodegeneration. Since its glyoxalase III activity catalyzing methylglyoxal (MG) to lactate was discovered, DJ-1 has been re-established as a deglycase decomposing the MG-intermediates with amino acids and nucleotides (hemithioacetal and hemiaminal) rather than MG itself, but it is still debatable. Here, we have clarified that human DJ-1 directly recognizes MG, and not MG-intermediates, by monitoring the detailed catalytic proces ...[more]

PMID: 37060572

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Project description:Highlights • Dicarbonyl metabolism in the mitochondria remains unclear.• Knockout mice for mitochondrial protein ES1 were generated.• Metabolic activity of glyoxal decreased significantly in ES1-knockout mice.• ES1 was found to have glyoxalase III activity and modulate dicarbonyl metabolism.• Mouse recombinant ES1 protein exhibited metabolic activity for glyoxal. Glycation, caused by reactive dicarbonyls, plays a role in various diseases by forming advanced glycation end products. In live cells, reactive dicarbonyls such as glyoxal (GO) and methylglyoxal (MGO) are produced during cell metabolism, and these should be removed consistently. However, the dicarbonyl metabolic system in the mitochondria remains unclear. It has been speculated that the mammalian mitochondrial protein ES1 is a homolog of bacterial elbB possessing glyoxalase III (GLO3) activity. Therefore, in this study, to investigate ES1 functions and GLO3 activity, we generated ES1-knockout (KO) mice and recombinant mouse ES1 protein and investigated the biochemical and histological analyses. In the mitochondrial fraction obtained from ES1-KO mouse brains, the GO metabolism and cytochrome c oxidase activity were significantly lower than those in the mitochondrial fraction obtained from wildtype (WT) mouse brains. However, the morphological features of the mitochondria did not change noticeably in the ES1-KO mouse brains compared with those in the WT mouse brains. The mitochondrial proteome analysis showed that the MGO degradation III pathway and oxidative phosphorylation-related proteins were increased. These should be the response to the reduced GO metabolism caused by ES1 deletion to compensate for the dicarbonyl metabolism and damaged cytochrome c oxidase by elevated GO. Recombinant mouse ES1 protein exhibited catalytic activity of converting GO to glycolic acid. These results indicate that ES1 possesses GLO3 activity and modulates the metabolism of GO in the mitochondria. To our knowledge, this is the first study to show a novel metabolic pathway for reactive dicarbonyls in mitochondria.

Dataset Information

Kinetic evidence in favor of glyoxalase III and against deglycase activity of DJ-1.

Publications

Kinetic evidence in favor of glyoxalase III and against deglycase activity of DJ-1.

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