Effects of maturation on the conformational free-energy landscape of SOD1

Abstract

Amyotrophic lateral sclerosis (ALS) is a devastating fatal syndrome characterized by very rapid degeneration of motor neurons. A leading hypothesis is that ALS is caused by toxic protein misfolding and aggregation, as also occurs in many other neurodegenerative disorders, such as prion, Alzheimer's, Parkinson's, and Huntington's diseases. A prominent cause of familial ALS is mutations in the protein superoxide dismutase (SOD1), which promote the formation of misfolded SOD1 conformers that are prone to aberrant interactions both with each other and with other cellular components. We have shown previously that immature SOD1, lacking bound Cu and Zn metal ions and the intrasubunit disulfide bond (apoSOD1), has a rugged free-energy surface (FES) and exchanges with four other conformations (excited states) that have millisecond lifetimes and sparse populations on the order of a few percent. Here, we examine further states of SOD1 along its maturation pathway, as well as those off-pathway resulting from metal loss that have been observed in proteinaceous inclusions. Metallation and disulfide bond formation lead to structural transformations including local ordering of the electrostatic loop and native dimerization that are observed in rare conformers of apoSOD1; thus, SOD1 maturation may occur via a population-switch mechanism whereby posttranslational modifications select for preexisting structures on the FES. Metallation and oxidation of SOD1 stabilize the native, mature conformation and decrease the number of detected excited conformational states, suggesting that it is the immature forms of the protein that contribute to misfolded conformations in vivo rather than the highly stable enzymatically active dimer.

Evolutionary Analyses of Sequence and Structure Space Unravel the Structural Facets of SOD1

“…Although there are multiple reports reflecting upon the importance of the electrostatic loops [21][22][23][24] in SOD1 and the role of metal ions in terms of crafting the structural integrity in SOD1, there is a lack of understanding as to how these residues are interdependent and whether they are under evolutionary selection pressure. Deploying co-evolution analysis on a wide array of sequences, we have been able to retrieve residues which are co-evolving and, hence, evolutionarily coupled.…”

Nonnative structure in a peptide model of the unfolded state of superoxide dismutase 1 (SOD1): Implications for ALS-linked aggregation

“…While the disulfide-intact, metal-bound SOD1 dimer is remarkably stable and resistant to aggregation (9), mutations are known to destabilize the native dimer and increase the populations of folded and unfolded forms of the monomeric protein that may nucleate aggregation (10)(11)(12)(13)(14). Several studies have suggested specific structures for such misfolded monomers and nonnative dimers (15)(16)(17)(18). However, the large number of ALS-linked SOD1 mutations and, in particular, their distribution throughout the sequence, motivated us to propose a general explanation for the aggregation phenomenon.…”

Peer Review #2 of "Methylene blue inhibits nucleation and elongation of SOD1 amyloid fibrils (v0.1)"

“…In this study, we have developed an assay allowing one to follow rapid and reproducible SOD1 kinetics at physiological pH (see Methods section). One key component used in the assay is a disulfide reducing agent, since disulfide reduction in SOD1 leads to destabilization of the SOD1 dimer and facilitates fibrillization (Furukawa, Torres & O'Halloran, 2004;Sheng et al, 2013;Culik et al, 2018). Another key component is guanidine hydrochloride (GuHCl), a chaotropic agent that accelerates SOD1 aggregation (Sheng et al, 2013) and in our case, strongly promotes SOD1 fibrillization and prevents "off-pathway" oligomer formation (Fig.…”