Werner-Allen Jonathan W, Monti Sarah, DuMond Jenna F, Levine Rodney L, Bax Ad
Biochemistry. 2018 Mar 6;57(9):1462-1474. doi: 10.1021/acs.biochem.7b01164. Epub 2018 Feb 15.
3,4-Dihydroxyphenylacetaldehyde (DOPAL) is a toxic and reactive product of dopamine catabolism. In the catecholaldehyde hypothesis for Parkinson's disease, it is a critical driver of the selective loss of dopaminergic neurons that characterizes the disease. DOPAL also cross-links α-synuclein, the main component of Lewy bodies, which are a pathological hallmark of the disease. We previously described the initial adduct formed in reactions between DOPAL and α-synuclein, a dicatechol pyrrole lysine (DCPL). Here, we examine the chemical basis for DOPAL-based cross-linking. We find that autoxidation of DCPL's catechol rings spurs its decomposition, yielding an intermediate dicatechol isoindole lysine (DCIL) product formed by an intramolecular reaction of the two catechol rings to give an unstable tetracyclic structure. DCIL then reacts with a second DCIL to give a dimeric, di-DCIL. This product is formed by an intermolecular carbon-carbon bond between the isoindole rings of the two DCILs that generates two structurally nonequivalent and separable atropisomers. Using α-synuclein, we demonstrate that the DOPAL-catalyzed formation of oligomers can be separated into two steps. The initial adduct formation occurs robustly within an hour, with DCPL as the main product, and the second step cross-links α-synuclein molecules. Exploiting this two-stage reaction, we use an isotopic labeling approach to show the predominant cross-linking mechanism is an interadduct reaction. Finally, we confirm that a mass consistent with a di-DCIL linkage can be observed in dimeric α-synuclein by mass spectrometry. Our work elucidates previously unknown pathways of catechol-based oxidative protein damage and will facilitate efforts to detect DOPAL-based cross-links in disease-state neurons.
3,4-二羟基苯乙醛（DOPAL）是多巴胺分解代谢产生的一种有毒且具有反应活性的产物。在帕金森病的儿茶酚醛假说中，它是该疾病特征性的多巴胺能神经元选择性丧失的关键驱动因素。DOPAL还能使路易小体的主要成分α-突触核蛋白发生交联，而路易小体是该疾病的一个病理标志。我们之前描述了DOPAL与α-突触核蛋白反应中形成的初始加合物，即二儿茶酚吡咯赖氨酸（DCPL）。在此，我们研究基于DOPAL交联的化学基础。我们发现DCPL儿茶酚环的自氧化促使其分解，产生一种中间产物二儿茶酚异吲哚赖氨酸（DCIL），它是由两个儿茶酚环通过分子内反应形成的不稳定四环结构。然后DCIL与另一个DCIL反应生成二聚体双DCIL。该产物是由两个DCIL的异吲哚环之间的分子间碳 - 碳键形成的，产生了两种结构上不等价且可分离的阻转异构体。使用α-突触核蛋白，我们证明DOPAL催化的寡聚体形成可分为两个步骤。初始加合物形成在一小时内强烈发生，主要产物为DCPL，第二步使α-突触核蛋白分子交联。利用这种两阶段反应，我们采用同位素标记方法表明主要的交联机制是加合物间反应。最后，我们通过质谱法证实，在二聚体α-突触核蛋白中可以观察到与双DCIL连接一致的质量。我们的工作阐明了基于儿茶酚的氧化蛋白损伤的先前未知途径，并将有助于在疾病状态的神经元中检测基于DOPAL的交联。
