Metal-free superoxide dismutase forms soluble oligomers under physiological conditions: A possible general mechanism for familial ALS

• Published in: Proceedings of the National Academy of Sciences - PNAS 2007-07, Vol.104 (27), p.11263-11267
• Main Authors: Banci, Lucia, Bertini, Ivano, Durazo, Armando, Girotto, Stefania, Gralla, Edith Butler, Martinelli, Manuele, Valentine, Joan Selverstone, Vieru, Miguela, Whitelegge, Julian P
• Format: Article
• Language: English
• Subjects: Aggregation; Amyloids; Amyotrophic lateral sclerosis; Amyotrophic Lateral Sclerosis - enzymology; Amyotrophic Lateral Sclerosis - genetics; Binding sites; Biological Sciences; Chromatography; Copper - chemistry; Copper - physiology; Dimers; Disulfides; Disulfides - chemistry; Fluorescence; Genetics; Humans; Metal ions; Mutation; Neurons; Oligomers; Protein folding; Saccharomyces cerevisiae - enzymology; Saccharomyces cerevisiae - genetics; Solubility; Superoxide Dismutase - chemistry; Superoxide Dismutase - genetics; Superoxide Dismutase - physiology; Superoxide Dismutase-1; Zinc; Zinc - chemistry; Zinc - physiology
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.0704307104

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disorder selectively affecting motor neurons; 90% of the total cases are sporadic, but 2% are associated with mutations in the gene coding for the antioxidant enzyme copper-zinc superoxide dismutase (SOD1). The causes of motor neuron death in ALS are poorly understood in general, but for SOD1-linked familial ALS, aberrant oligomerization of SOD1 mutant proteins has been strongly implicated. In this work, we show that wild-type human SOD1, when lacking both its metal ions, forms large, stable, soluble protein oligomers with an average molecular mass of [almost equal to]650 kDa under physiological conditions, i.e., 37°C, pH 7.0, and 100 μM protein concentration. It further is shown here that intermolecular disulfide bonds are formed during oligomerization and that Cys-6 and Cys-111 are implicated in this bonding. The formation of the soluble oligomers was monitored by their ability to enhance the fluorescence of thioflavin T, a benzothiazole dye that increases in fluorescence intensity upon binding to amyloid fibers, and by disruption of this binding upon addition of the chaotropic agent guanidine hydrochloride. Our results suggest a general, unifying picture of SOD1 aggregation that could operate when wild-type or mutant SOD1 proteins lack their metal ions. Although we cannot exclude other mechanisms in SOD1-linked familial ALS, the one proposed here has the strength of explaining how a large and diverse set of SOD1 mutant proteins all could lead to disease through the same mechanism.