McDowall Jennifer S, Ntai Ioanna, Hake Jonathon, Whitley Paul R, Mason Jody M, Pudney Christopher R, Brown David R
Department of Biology and Biochemistry, Faculty of Science, University of Bath , Bath, U.K.
Biochemistry. 2017 May 16;56(19):2497-2505. doi: 10.1021/acs.biochem.7b00257. Epub 2017 May 4.
α-Synuclein (α-syn) is a cytosolic protein known for its association with neurodegenerative diseases, including Parkinson's disease and other synucleinopathies. The potential cellular function of α-synuclein may be of consequence for understanding the pathogenesis of such diseases. Previous work has suggested that α-synuclein can catalyze the reduction of iron as a ferrireductase. We performed a detailed analysis of the steady-state kinetics of recombinant α-syn ferrireductase activity and for disease-associated variants. Our study illustrates that the ferrireductase activity we observed is clearly commensurate with bona fide enzyme activity and suggests a mechanistic rationale for the activity and the relationship to cellular regulation of the pool of Fe(III) and Fe(II). Using cell-based studies, we examined the functionally active conformation and found that the major catalytically active form is a putative membrane-associated tetramer. Using an artificial membrane environment with recombinant protein, we demonstrate that secondary structure folding of α-synuclein is insufficient to allow enzyme activity and the absolute specificity of the tertiary/quaternary structure is the primary requirement. Finally, we explored the steady-state kinetics of a range of disease α-synuclein variants and found that variants involved in neurodegenerative disease exhibited major changes in their enzymatic activity. We discuss these data in the context of a potential disease-associated mechanism for aberrant α-synuclein ferrireductase activity.
α-突触核蛋白（α-syn）是一种胞质蛋白，因其与神经退行性疾病（包括帕金森病和其他突触核蛋白病）的关联而闻名。α-突触核蛋白的潜在细胞功能可能对理解此类疾病的发病机制具有重要意义。先前的研究表明，α-突触核蛋白可以作为铁还原酶催化铁的还原。我们对重组α-突触核蛋白铁还原酶活性及其疾病相关变体的稳态动力学进行了详细分析。我们的研究表明，我们观察到的铁还原酶活性显然与真正的酶活性相当，并提出了该活性及其与Fe(III)和Fe(II)池细胞调节关系的机制原理。通过基于细胞的研究，我们研究了功能活性构象，发现主要的催化活性形式是一种假定的膜相关四聚体。使用含有重组蛋白的人工膜环境，我们证明α-突触核蛋白的二级结构折叠不足以产生酶活性，三级/四级结构的绝对特异性是主要要求。最后，我们研究了一系列疾病α-突触核蛋白变体的稳态动力学，发现与神经退行性疾病相关的变体在酶活性方面表现出重大变化。我们在异常α-突触核蛋白铁还原酶活性的潜在疾病相关机制的背景下讨论了这些数据。
