Hulleman John D, Mirzaei Hamid, Guigard Emmanuel, Taylor Kellie L, Ray Soumya S, Kay Cyril M, Regnier Fred E, Rochet Jean-Christophe
Department of Medicinal Chemistry and Molecular Pharmacology, Purdue University, West Lafayette, Indiana 47907, USA.
Biochemistry. 2007 May 15;46(19):5776-89. doi: 10.1021/bi7001778. Epub 2007 Apr 24.
Parkinson's disease (PD) is a neurodegenerative disorder characterized by oxidative stress and protein aggregation. Both toxic phenomena are mitigated by DJ-1, a homodimeric protein with proposed antioxidant and chaperone activities. The neuroprotective function of DJ-1 is modulated by oxidation of cysteine 106, a residue that may act as an oxidative stress sensor. Loss-of-function mutations in the DJ-1 gene have been linked to early onset PD, and age-dependent over-oxidation of DJ-1 is thought to contribute to sporadic PD. The familial mutant L166P fails to dimerize and is rapidly degraded, suggesting that protein destabilization accounts for the dysfunction of this mutant. In this study, we investigated how the structure and stability of DJ-1 are impacted by two other pathogenic substitutions (M26I and E64D) and by over-oxidation with H2O2. Whereas the recombinant wild-type protein and E64D both adopted a stable dimeric structure, M26I showed an increased propensity to aggregate and decreased secondary structure. Similar to M26I, over-oxidized wild-type DJ-1 exhibited reduced secondary structure, and this property correlated with destabilization of the dimer. The engineered mutant C106A had a greater thermodynamic stability and was more resistant to oxidation-induced destabilization than the wild-type protein. These results suggest that (i) the M26I substitution and over-oxidation destabilize dimeric DJ-1, and (ii) the oxidation of cysteine 106 contributes to DJ-1 destabilization. Our findings provide a structural basis for DJ-1 dysfunction in familial and sporadic PD, and they suggest that dimer stabilization is a reasonable therapeutic strategy to treat both forms of this disorder.
帕金森病（PD）是一种以氧化应激和蛋白质聚集为特征的神经退行性疾病。DJ-1可减轻这两种毒性现象，它是一种同二聚体蛋白，具有抗氧化和伴侣活性。DJ-1的神经保护功能受半胱氨酸106氧化的调节，该残基可能作为氧化应激传感器。DJ-1基因的功能丧失突变与早发性帕金森病有关，DJ-1的年龄依赖性过度氧化被认为与散发性帕金森病有关。家族性突变体L166P无法形成二聚体并迅速降解，这表明蛋白质不稳定是该突变体功能障碍的原因。在本研究中，我们研究了另外两种致病取代（M26I和E64D）以及H2O2过度氧化如何影响DJ-1的结构和稳定性。重组野生型蛋白和E64D均采用稳定的二聚体结构，而M26I表现出更高的聚集倾向和二级结构减少。与M26I类似，过度氧化的野生型DJ-1二级结构减少，且该特性与二聚体不稳定相关。工程突变体C106A具有更高的热力学稳定性，比野生型蛋白更能抵抗氧化诱导的不稳定。这些结果表明：（i）M26I取代和过度氧化使二聚体DJ-1不稳定；（ii）半胱氨酸106的氧化导致DJ-1不稳定。我们的研究结果为家族性和散发性帕金森病中DJ-1功能障碍提供了结构基础，并表明二聚体稳定化是治疗这两种形式疾病的合理治疗策略。
