Department of Chemistry and Biochemistry, and ‡Institute of Biomedical Studies, Baylor University , Waco, Texas 76798-7348, United States.
ACS Chem Neurosci. 2016 Jun 15;7(6):799-810. doi: 10.1021/acschemneuro.6b00048. Epub 2016 Mar 31.
Recent reports suggest that the nucleation and propagation of oligomeric superoxide dismutase-1 (SOD1) is effectively stochastic in vivo and in vitro. This perplexing kinetic variability-observed for other proteins and frequently attributed to experimental error-plagues attempts to discern how SOD1 mutations and post-translational modifications linked to amyotrophic lateral sclerosis (ALS) affect SOD1 aggregation. This study used microplate fluorescence spectroscopy and dynamic light scattering to measure rates of fibrillar and amorphous SOD1 aggregation at high iteration (ntotal = 1.2 × 10(3)). Rates of oligomerization were intrinsically irreproducible and populated continuous probability distributions. Modifying reaction conditions to mimic random and systematic experimental error could not account for kinetic outliers in standard assays, suggesting that stochasticity is not an experimental artifact, rather an intrinsic property of SOD1 oligomerization (presumably caused by competing pathways of oligomerization). Moreover, mean rates of fibrillar and amorphous nucleation were not uniformly increased by mutations that cause ALS; however, mutations did increase kinetic noise (variation) associated with nucleation and propagation. The stochastic aggregation of SOD1 provides a plausible statistical framework to rationalize how a pathogenic mutation can increase the probability of oligomer nucleation within a single cell, without increasing the mean rate of nucleation across an entire population of cells.
最近的报告表明，寡聚超氧化物歧化酶-1（SOD1）的成核和增殖在体内和体外都是有效的随机过程。这种令人困惑的动力学可变性——在其他蛋白质中也经常观察到，并常归因于实验误差——阻碍了人们试图辨别与肌萎缩侧索硬化症（ALS）相关的 SOD1 突变和翻译后修饰如何影响 SOD1 聚集。本研究使用微孔板荧光光谱法和动态光散射法在高迭代（ntotal = 1.2 × 10(3)）下测量纤维状和无定形 SOD1 聚集的速率。低聚物的聚合速率本质上是不可重复的，并呈现连续的概率分布。模拟随机和系统实验误差的反应条件的改变不能解释标准测定中的动力学异常值，这表明随机性不是实验误差，而是 SOD1 低聚物聚合的固有特性（可能是由低聚物聚合的竞争途径引起的）。此外，导致 ALS 的突变并没有均匀地增加纤维状和无定形核形成的平均速率；然而，突变确实增加了与成核和增殖相关的动力学噪声（变化）。SOD1 的随机聚集提供了一个合理的统计框架，可以解释致病性突变如何在单个细胞内增加寡聚物成核的概率，而不会增加整个细胞群体中成核的平均速率。
