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

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 approximately 650 kDa under physiological conditions, i.e., 37 degrees C, pH 7.0, and 100 microM 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.