Abstract
Dominant mutations in Cu,Zn-superoxide dismutase (SOD1) cause a familial form of amyotrophic lateral sclerosis (fALS). Misfolding and aggregation of mutant SOD1 proteins are a pathological hallmark of SOD1-related fALS cases; however, the molecular mechanism of SOD1 aggregation remains controversial. Here, I have used E. coli as a model organism and shown multiple distinct pathways of SOD1 aggregation that are dependent upon its thiol-disulfide status. Overexpression of fALS-mutant SOD1s in the cytoplasm of E. coli BL21 and SHuffleTM, where redox environment is reducing and oxidizing, respectively, resulted in the formation of insoluble aggregates with notable differences; a disulfide bond of SOD1 was completely reduced in BL21 or abnormally formed between SOD1 molecules in SHuffleTM. Depending upon intracellular redox environment, therefore, mutant SOD1 is considered to misfold/aggregate through distinct pathways, which would be relevant in description of the pathological heterogeneity of SOD1-related fALS cases.
Highlights
Thiol-disulfide status is critical for functioning of many proteins (Sevier and Kaiser, 2002), and Cu,Zn-superoxide dismutase (SOD1) is one of such proteins in which formation of an intramolecular disulfide bond is required for folding into its enzymatically active conformation (Furukawa et al, 2004)
Abnormalities in a thiol-disulfide status of SOD1 have been proposed as a pathological change in a familial form of amyotrophic lateral sclerosis that is caused by dominant mutations in SOD1 (Rosen et al, 1993)
SOD1 AGGREGATES UNDER REDUCING ENVIRONMENT OF E. coli CYTOPLASM Introduction of an intramolecular disulfide bond between Cys57 and Cys146 in SOD1 has been known to increase the electrophoretic mobility of SOD1 (Furukawa et al, 2004); thiol-disulfide status of SOD1 can be determined by non-reducing SDS-PAGE analysis
Summary
Thiol-disulfide status is critical for functioning of many proteins (Sevier and Kaiser, 2002), and Cu,Zn-superoxide dismutase (SOD1) is one of such proteins in which formation of an intramolecular disulfide bond is required for folding into its enzymatically active conformation (Furukawa et al, 2004). In transgenic mice expressing human SOD1 with fALS mutations, mutant SOD1 has been shown to form insoluble oligomers cross-linked via intermolecular disulfide bonds (Furukawa et al, 2006). FALS-mutant human SOD1 proteins in E. coli BL21 were found to exist as a disulfide-reduced state and form insoluble fibrillar aggregates.
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