Karch Celeste M, Borchelt David R
Department of Neuroscience, McKnight Brain Institute, SantaFe HealthCare Alzheimer's Disease Research Center, University of Florida, Gainesville, Florida 32611, USA.
J Biol Chem. 2008 May 16;283(20):13528-37. doi: 10.1074/jbc.M800564200. Epub 2008 Mar 3.
One of the mechanisms by which mutations in superoxide dismutase 1 (SOD1) cause familial amyotrophic lateral sclerosis (fALS) is proposed to involve the accumulation of detergent-insoluble, disulfide-cross-linked, mutant protein. Recent studies have implicated cysteine residues at positions 6 and 111 as critical in mediating disulfide cross-linking and promoting aggregation. In the present study, we used a panel of experimental and disease-linked mutations at cysteine residues of SOD1 (positions 6, 57, 111, and 146) in cell culture assays for aggregation to demonstrate that extensive disulfide cross-linking is not required for the formation of mutant SOD1 aggregates. Experimental mutants possessing only a single cysteine residue or lacking cysteine entirely were found to retain high potential to aggregate. Furthermore we demonstrate that aggregate structures in symptomatic SOD1-G93A mice can be dissociated such that they no longer sediment upon ultracentrifugation (i.e. appear soluble) under relatively mild conditions that leave disulfide bonds intact. Similar to other recent work, we found that cysteines 6 and 111, particularly the latter, play interesting roles in modulating the aggregation of human SOD1. However, we did not find that extensive disulfide cross-linking via these residues, or any other cysteine, is critical to aggregate structure. Instead we suggest that these residues participate in other features of the protein that, in some manner, modulate aggregation.
超氧化物歧化酶1（SOD1）突变导致家族性肌萎缩侧索硬化症（fALS）的一种机制被认为涉及去污剂不溶性、二硫键交联的突变蛋白的积累。最近的研究表明，第6位和第111位的半胱氨酸残基在介导二硫键交联和促进聚集方面至关重要。在本研究中，我们在细胞培养聚集试验中使用了一组位于SOD1半胱氨酸残基（第6、57、111和146位）的实验性和疾病相关突变，以证明突变型SOD1聚集体的形成不需要广泛的二硫键交联。发现仅具有单个半胱氨酸残基或完全缺乏半胱氨酸的实验性突变体仍保留高聚集潜力。此外，我们证明，有症状的SOD1 - G93A小鼠中的聚集体结构可以解离，使得它们在保持二硫键完整的相对温和条件下超速离心时不再沉淀（即看起来是可溶的）。与其他近期研究相似，我们发现第6位和第111位的半胱氨酸，尤其是后者，在调节人SOD1的聚集方面发挥着有趣的作用。然而，我们没有发现通过这些残基或任何其他半胱氨酸进行的广泛二硫键交联对聚集体结构至关重要。相反，我们认为这些残基参与了蛋白质的其他特征，这些特征以某种方式调节聚集。
