Abstract：
Mutations in the protein superoxide dismutase-1 (SOD1) promote its misfolding and aggregation, ultimately causing familial forms of the debilitating neurodegenerative disease amyotrophic lateral sclerosis (ALS). Currently, over 220 (mostly missense) ALS-causing mutations in the SOD1 protein have been identified, indicating that common structural features are responsible for aggregation and toxicity. Using in silico tools, we predicted amyloidogenic regions in the ALS-associated SOD1-G85R mutant, finding seven regions throughout the structure. Introduction of proline residues into β-strands II (I18P) or III (I35P) reduced the aggregation propensity and toxicity of SOD1-G85R in cells, significantly more so than proline mutations in other amyloidogenic regions. The I18P and I35P mutations also reduced the capability of SOD1-G85R to template onto previously formed non-proline mutant SOD1 aggregates as measured by fluorescence recovery after photobleaching. Finally, we found that, while the I18P and I35P mutants are less structurally stable than SOD1-G85R, the proline mutants are less aggregation-prone during proteasome inhibition, and less toxic to cells overall. Our research highlights the importance of a previously underappreciated SOD1 amyloidogenic region in β-strand II (15QGIINF20) to the aggregation and toxicity of SOD1 in ALS mutants, and suggests that β-strands II and III may be good targets for the development of SOD1-associated ALS therapies.
摘要：
蛋白质超氧化物歧化酶-1（SOD1）的突变促进其错误折叠和聚集，最终导致家族性形式的衰弱性神经退行性疾病肌萎缩侧索硬化症（ALS）。目前,已鉴定出超过220个（主要是错义）引起SOD1蛋白的ALS突变，表明共同的结构特征是聚集和毒性的原因。使用硅片工具，我们预测了ALS相关SOD1-G85R突变体中的淀粉样区域，在整个结构中找到七个区域。将脯氨酸残基引入β链II（I18P）或III（I35P）降低了SOD1-G85R在细胞中的聚集倾向和毒性，明显高于其他淀粉样蛋白区域的脯氨酸突变。I18P和I35P突变还降低了SOD1-G85R模板到先前形成的非脯氨酸突变体SOD1聚集体上的能力，如通过光漂白后的荧光恢复所测量的。最后,我们发现,虽然I18P和I35P突变体的结构稳定性不如SOD1-G85R，脯氨酸突变体在蛋白酶体抑制期间不易聚集，对整体细胞毒性较小。我们的研究强调了β链II（15QGIINF20）中先前未被重视的SOD1淀粉样区对ALS突变体中SOD1的聚集和毒性的重要性，并表明β链II和III可能是开发SOD1相关ALS疗法的良好靶标。
