The Disulfide Bond, but Not Zinc or Dimerization, Controls Initiation and Seeded Growth in Amyotrophic Lateral Sclerosis-linked Cu,Zn Superoxide Dismutase (SOD1) Fibrillation

Abstract

Aggregation of copper-zinc superoxide dismutase (SOD1) is a defining feature of familial ALS caused by inherited mutations in the sod1 gene, and misfolded and aggregated forms of wildtype SOD1 are found in both sporadic and familial ALS cases. Mature SOD1 owes its exceptional stability to a number of posttranslational modifications as follows: formation of the intramolecular disulfide bond, binding of copper and zinc, and dimerization. Loss of stability due to the failure to acquire one or more of these modifications is proposed to lead to aggregation in vivo. Previously, we showed that the presence of apo-, disulfide-reduced SOD1, the most immature form of SOD1, results in initiation of fibrillation of more mature forms that have an intact Cys-57-Cys-146 disulfide bond and are partially metallated. In this study, we examine the ability of each of the above post-translational modifications to modulate fibril initiation and seeded growth. Cobalt or zinc binding, despite conferring great structural stability, neither inhibits the initiation propensity of disulfide-reduced SOD1 nor consistently protects disulfide-oxidized SOD1 from being recruited into growing fibrils across wild-type and a number of ALS mutants. In contrast, reduction of the disulfide bond, known to be necessary for fibril initiation, also allows for faster recruitment during seeded amyloid growth. These results identify separate factors that differently influence seeded growth and initiation and indicate a lack of correlation between the overall thermodynamic stability of partially mature SOD1 states and their ability to initiate fibrillation or be recruited by a growing fibril. Familial amyotrophic lateral sclerosis (FALS)3 caused by mutations in the SOD1 gene shows similarities to other members of the large class of neurodegenerative disorders that are characterized by gradual aggregation of specific proteins and subsequent cell death in characteristic regions of the central nervous system. Some prominent examples are amyloid-␤ deposits in Alzheimer disease, ␣-synuclein and hyperphosphorylated Tau protein deposits in Parkinson disease, prion protein deposits in the transmissible spongiform encephalopathies, and huntingtin deposits in Huntington disease (1). In the case of SOD1-linked FALS, protein deposits have been identified in affected tissues from patients and from ALS-SOD1 transgenic mice consisting largely of aggregated copper,zinc superoxide dismutase protein (Cu,Zn-SOD, SOD1 protein).Our current understanding of this class of diseases is that the implicated proteins misfold, aggregate, and ultimately deposit as extracellular plaques or intracellular inclusions in the afflicted regions of the CNS. Although the exact structures contributing to these heterogeneous protein aggregates are unknown, such diseases are nevertheless usually referred to as "amyloid diseases" because fibrillar properties characteristic of cross-␤-sheet amyloid are often detected in them and because each of the isolated proteins implicated in causing disease shows a...

Cellular Redox Systems Impact the Aggregation of Cu,Zn Superoxide Dismutase Linked to Familial Amyotrophic Lateral Sclerosis

“…WT hSOD1 is much more resistant to reduction by both systems. Disulfide reduction and metal ion removal may trigger SOD1 aggregation (42), but, in fact, disulfide bonds cause the initiation of its fibrillation (43). In our study, the reduced mutant hSOD1 was detected to be catalytically active.…”

Misfolded SOD1 is not a primary component of sporadic ALS

“…It has been reported that certain posttranslational modifications of wild-type SOD1 (SOD1 WT ), including oxidation [6, 12, 16, 26, 27, 37] and demetallation [47] induce its misfolding in vitro , suggesting that even SOD1 WT can acquire toxic properties like those associated with ALS-linked mutants of SOD1. In transgenic mice, ~5–20 fold higher levels of SOD1 WT expression cause SOD1 aggregation and neuronal pathology in the CNS in aged mice [24, 35, 36], whereas expressing low levels of human SOD1 WT does not result in ALS-like disease [9, 14, 28].…”

A Phosphomimetic Mutation Stabilizes SOD1 and Rescues Cell Viability in the Context of an ALS-Associated Mutation

“…In familial and sporadic ALS cases, post-translational modifications induced by environmental factors play a significant role in disease progression (Barber and Shaw, 2010; Chattopadhyay et al, 2015; Pasinelli and Brown, 2006). Specifically, glutathionylation of C111, oxidative modification of W32, and phosphorylation of T2 or S58/T59 have been identified in human SOD1 (Coelho et al, 2014; Wilcox et al, 2009).…”