The impact of post-translational modifications on aggregation of Cu, Zn superoxide dismutase in amyotrophic lateral sclerosis

Redler, Rachel L.

The impact of post-translational modifications on aggregation of Cu, Zn superoxide dismutase in amyotrophic lateral sclerosis

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March 20, 2019

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Redler, Rachel L.
- Affiliation: School of Medicine, Department of Biochemistry and Biophysics

Abstract

Aberrant conformers of disease-linked proteins have been proposed as cytotoxic agents in several late-onset neurodegenerative disorders, including Alzheimer's disease and amyotrophic lateral sclerosis (ALS). Mutations in the gene encoding Cu, Zn superoxide dismutase (SOD1) are present in a subset of familial ALS (FALS) cases; most of these mutations destabilize the protein, although typically by a small margin relative to SOD1's exceptionally high stability. Therefore, SOD1 with FALS-linked substitutions often misfolds and aggregates, adopting aberrant conformations that interact with numerous cellular components and disrupt their functioning, despite having a more stable folded state than would be expected for an aggregation-prone protein. This fact, together with the specific death of motor neurons late in life despite ubiquitous expression of mutant SOD1 since birth, implicates factors in the cellular environment as substantial contributors to the cytotoxicity of mutant SOD1 in FALS. One non-genetic factor likely to influence misfolding and aggregation of SOD1 in human tissue is its susceptibility to abundant post-translational modifications, including phosphorylation and numerous oxidative modifications. We find that reversible oxidative modification of Cys-111 by the glutathione tripeptide destabilizes the native SOD1 homodimer, increasing the equilibrium dissociation constant of the WT dimer from low nanomolar to the low micromolar range, and further destabilizes SOD1 containing a FALS-linked substitution within the dimer interface (A4V). Assessment of the effect of glutathionylation on dimer dissociation kinetics using surface plasmon resonance revealed that this modification causes minimal change in dimer dissociation rate; therefore, the increased Kd observed for glutathionylated WT and A4V SOD1 must result from slowed association of modified monomers. In addition to inducing dissociation of the native dimer, Cys-111 glutathionylation promotes the assembly of soluble non-native oligomers that contain an epitope specific to disease-relevant misfolded SOD1. Our findings suggest that soluble non-native SOD1 oligomers share structural similarity to pathogenic misfolded species found in ALS patients, and therefore represent potential cytotoxic agents and therapeutic targets in ALS. Furthermore, the induction of SOD1 misfolding and aggregation by glutathionylation represents a possible mechanism by which oxidative stress brought on by aging triggers the transition of SOD1 from its natively folded state to cytotoxic conformations.

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