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Abstract：
DJ-1 is a multifunctional protein associated with cancers and autosomal early-onset Parkinson disease. Besides the well-documented antioxidative stress activity, recent studies show that DJ-1 has deglycation enzymatic activity and anti-ferroptosis effect. It has been shown that DJ-1 forms the homodimerization, which dictates its antioxidative stress activity. In this study, we investigated the relationship between the dimeric structure of DJ-1 and its newly reported activities. In HEK293T cells with Flag-tagged and Myc-tagged DJ-1 overexpression, we performed deletion mutations and point mutations, narrowed down the most critical motif at the C terminus. We found that the deletion mutation of the last three amino acids at the C terminus of DJ-1 (DJ-1 ΔC3) disrupted its homodimerization with the hydrophobic L187 residue being of great importance for DJ-1 homodimerization. In addition, the ability in methylglyoxal (MGO) detoxification and deglycation was almost abolished in the mutation of DJ-1 ΔC3 and point mutant L187E compared with wild-type DJ-1 (DJ-1 WT). We also showed the suppression of erastin-triggered ferroptosis in DJ-1-/- mouse embryonic fibroblast cells was abolished by ΔC3 and L187E, but partially diminished by V51C. Thus, our results demonstrate that the C terminus of DJ-1 is crucial for its homodimerization, deglycation activity, and suppression of ferroptosis.
摘要：
DJ-1是一种与癌症和常染色体早发性帕金森病相关的多功能蛋白。除了有据可查的抗氧化应激活性，近年来研究表明,DJ-1具有去糖基化酶活性和抗铁凋亡作用。研究表明，DJ-1形成了同源二聚化，这决定了它的抗氧化应激活性。在这项研究中,我们研究了DJ-1的二聚体结构与其新报道的活性之间的关系。在具有Flag标记和Myc标记的DJ-1过表达的HEK293T细胞中，我们进行了缺失突变和点突变，缩小了C末端最关键的主题。我们发现DJ-1（DJ-1ΔC3）C末端最后三个氨基酸的缺失突变破坏了其同源二聚化，疏水性L187残基对DJ-1同源二聚化至关重要。此外,与野生型DJ-1（DJ-1WT）相比，DJ-1ΔC3和点突变体L187E的突变几乎消除了甲基乙二醛（MGO）解毒和去糖基化的能力。我们还显示，ΔC3和L187E消除了DJ-1-/-小鼠胚胎成纤维细胞中擦除素触发的铁凋亡的抑制，但部分减少了V51C。因此，我们的结果表明，DJ-1的C末端对其同二聚化至关重要，去糖基化活性，和铁性凋亡的抑制。
