Abstract
Dominant mutations in Cu,Zn-superoxide dismutase (SOD1) are a cause of a familial form of amyotrophic lateral sclerosis. Wild-type SOD1 forms a highly conserved intra-molecular disulfide bond, whereas pathological SOD1 proteins are cross-linked via intermolecular disulfide bonds and form insoluble oligomers. A thiol-disulfide status in SOD1 will thus play a regulatory role in determining its folding/misfolding pathways; however, it remains unknown how pathogenic mutations in SOD1 affect the thiol-disulfide status to facilitate the protein misfolding. Here, we show that the structural destabilization of SOD1 scrambles a disulfide bond among four Cys residues in an SOD1 molecule. The disulfide scrambling produces SOD1 monomers with distinct electrophoretic mobility and also reproduces the formation of disulfide-linked oligomers. We have also found that the familial form of amyotrophic lateral sclerosis-causing mutations facilitate the disulfide scrambling in SOD1. Based upon our results, therefore, scrambling of the conserved disulfide bond will be a key event to cause the pathological changes in disease-associated mutant SOD1 proteins.
Highlights
Cu,Zn-superoxide dismutase (SOD1) possesses a highly conserved intramolecular disulfide bond
We have found that the familial form of amyotrophic lateral sclerosis-causing mutations facilitate the disulfide scrambling in SOD1
Distinct Aggregation Pathways of Apo-SOD1 with and without a Conserved Disulfide Bond—An enzymatically active form of SOD1 is equipped with copper and zinc ions and an intramolecular disulfide bond [6]
Summary
Cu,Zn-superoxide dismutase (SOD1) possesses a highly conserved intramolecular disulfide bond. Results: Structural destabilization of SOD1 scrambles the intramolecular disulfide to form cross-linked oligomers with an intermolecular disulfide bond. Based upon our results scrambling of the conserved disulfide bond will be a key event to cause the pathological changes in disease-associated mutant SOD1 proteins. Significant amounts of SOD1 that appears to be in the disulfide-reduced state have been reported to constitute the insoluble aggregates observed in fALS model mice [11, 14], where non-native intermolecular disulfide bonds have been further found to cross-link the SOD1 oligomers [14, 16, 17]. Structural destabilization of the apo-SOD1 with a canonical intramolecular disulfide bond (Cys57–Cys146) triggers the nucleophilic attack of Cys6/111 to the disulfide bond between Cys and Cys146 Such disulfide isomerization first occurs within an SOD1 molecule and proceeds between the molecules to form insoluble disulfide-linked oligomers. We propose that the disulfide reduction is not always required for SOD1 misfolding; rather, the disulfide scrambling by intra- and intermolecular isomerization constitutes a relevant pathway for the aggregation of mutant SOD1 proteins
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