Background/Objectives: Approximately 20% of familial ALS (fALS) cases are linked to mutations in Cu/Zn superoxide dismutase (SOD1). Through a gain function, SOD1 misfolding exerts a toxic effect on motor neurons, leading to their degradation and ALS symptomology in both fALS cases and
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Background/Objectives: Approximately 20% of familial ALS (fALS) cases are linked to mutations in Cu/Zn superoxide dismutase (SOD1). Through a gain function, SOD1 misfolding exerts a toxic effect on motor neurons, leading to their degradation and ALS symptomology in both fALS cases and sporadic ALS (sALS) cases with no known genetic cause. To further our understanding of SOD1-ALS etiology, identifying motor neuron-specific SOD1 post-translational modifications (PTMs) and studying their structural influence is necessary. To this end, we have conducted a study on the influence of the deamidation of Asn53, a PTM proximal to key stabilizing motifs in SOD1, which has scarcely been addressed in the literature to date.
Methods: Deamidation to N53 was identified by tandem mass spectrometry of SOD1 immunoprecipitated from motor neuron (MN) cultures derived from wild-type (WT) human induced pluripotent stem cells (iPSCs). WT SOD1 and N53D SOD1, a mutant mimicking the deamidation, were expressed in
Escherichia coli and purified for in vitro analyses. Differences between species were measured by experiments probing metal cofactors, relative monomer populations, and aggregation propensity. Furthermore, molecular dynamics experiments were conducted to model and determine the influence of the PTM on SOD1 structure.
Results: In contrast to WT, N53D SOD1 showed non-native incorporation of metal cofactors, coordinating more Zn
2+ cofactors than total Zn-binding sites, and more readily adopted monomeric forms, unfolded, and aggregated with heating, possibly while releasing coordinated metals.
Conclusions: Deamidation to N53 in SOD1 encourages the adoption of non-native conformers, and its detection in WT MN cultures suggests relevance to sALS pathophysiology.
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