Amyotrophic Lateral Sclerosis-associated Copper/Zinc Superoxide Dismutase Mutations Preferentially Reduce the Repulsive Charge of the Proteins

Abstract

We provide bioinformatical evidence that protein charge plays a key role in the disease mechanism of amyotrophic lateral sclerosis (ALS). Analysis of 100 ALS-associated mutations in copper/zinc superoxide dismutase (SOD1) shows that these are site-selective with a preference to decrease the proteins' net repulsive charge. For each SOD1 monomer this charge is normally ؊6. Because biomolecules as a rule maintain net negative charge to assure solubility in the cellular interior, the result lends support to the hypothesis of protein aggregation as an initiating event in the ALS pathogenesis. The strength of the preferential reduction of repulsive charge is higher in SOD1-associated ALS than in other inherited protein disorders.The motor-neuron disorder amyotrophic lateral sclerosis (ALS) 2 has been associated to more than 100 missense mutations in the radical scavenger copper/zinc superoxide dismutase (SOD1) (1). In analogy with several other neurodegenerative disorders (2), ALS seems associated with protein misfolding and aggregation (3-8). The principal support for such a disease mechanism is that the neural damage is accompanied by the accumulation of intracellular SOD1 deposits (9, 10). Even so, it is not the mature protein inclusions that seem to pose the main problem but rather some yet unidentified precursor accumulating in connection to the macroscopic aggregation process (11)(12)(13)(14)(15)(16). Consistent with such a scenario, transgenic mice suffer neuron stress long before any SOD1 inclusions can be detected (10,17,18). At the molecular level, studies of how the individual ALS-associated mutations affect the SOD1 molecule have further indicated protein stability as an important factor in the disease mechanism (19 -25); the more severe the destabilization caused by the mutation, the faster the disease progression (20). In this study, we shed further light on the disease-provoking properties of the SOD1 molecule by examining bioinformatically the chemical nature of 100 ALS-associated missense mutations. The results reveal an additional disease factor; the mutations have an overall tendency to decrease the net negative charge of the proteins. The negative charge repulsion that generally assures macromolecular solubility in vivo is diminished. This tendency is stronger for ALS mutations than in other hereditary protein disorders and cannot be discerned in the interspecies variability of the SOD1 sequence. In the few contrasting cases where the SOD1 mutations produce an increased repulsive charge, the side chain substitution tends also to be structurally obstructive and to occur in strictly conserved positions. That is, the opposed effect of increased repulsive charge is not sufficiently strong to compensate for the accompanying penalties in protein stability. The observation shows that the decreased net negative charge of SOD1 is a critical contributory factor to the ability of the mutation to cause neural damage. Taken together with the impact of decreased protein stability, the data also provide genetic s...

Decreased stability and increased formation of soluble aggregates by immature superoxide dismutase do not account for disease severity in ALS

“…Correlations between the properties of mutant SOD1 and ALS disease characteristics have been sought for many years and are critical for deciphering disease mechanisms. Previous studies have reported evidence for a weak inverse correlation between oxidized apo SOD1 stability and ALS disease duration (45)(46)(47), which improves when global or local protein characteristics such as charge (48) or hydrogen bonding (45) are considered. The results for V148I suggest that increased hydrophobicity of the exposed dimer interface may be another significant modulator of aggregation.…”

Transient structural distortion of metal-free Cu/Zn superoxide dismutase triggers aberrant oligomerization

“…Our data reveal molecular properties of SOD1 that may offer insight into the variations in disease propensity and progression observed among different mutants. Previous reports have shown that the net electric charge of SOD1 mutants is an important factor in ALS (14,28). Because the population of monomeric apo-SOD1 depends on a number of factors, including dimer interface integrity, metal affinity, and disulfide oxidation state, mutations compromising any of these properties would be expected to influence disease propensity (2, 11).…”

“…The monomeric apo states of A4V, G85R, and G93A are all highly destabilized toward global unfolding, whereas D90A imparts only a marginal effect in this respect (14). G85R and D90A have a lower net negative charge, which increases the aggregation propensity (14,28), and G85R has a lower metal affinity, which increases the relative population of apo-SOD1 (2). A4V and G93A confer short survival times in patients with ALS (Ͻ 3 years), while G85R and D90A are associated with more variable disease progression (1,2).…”

“…More than 100 different mutations in SOD1 have been linked to familial ALS, but how the great variation in molecular properties (e.g., stability and metal affinity) among these mutants relate to the same disease is not well understood (2,28). To investigate this issue, we studied both WT SOD1 and 4 familial ALS mutants.…”

Genome-wide association analysis reveals a SOD1 mutation in canine degenerative myelopathy that resembles amyotrophic lateral sclerosis

“…These 4 dogs may have been misdiagnosed. The SOD1:c.118GϾA transition predicts an E40K missense with anionic membrane surfaces (10,25). A glutamate at a position corresponding to amino acid position 40 in canine SOD1 is conserved in 19 of 20 mammals identified in a Blastp query of the nonredundant protein sequences in the National Center for Biotechnology Information database (Fig.…”