Intermolecular transmission of superoxide dismutase 1 misfolding in living cells

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DOIResolve DOI: http://doi.org/10.1073/pnas.1102645108
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TypeArticle
Journal titleProceedings of the National Academy of Sciences of the United States of America
ISSN0027-8424
Volume108
Issue39
Pages1639816403; # of pages: 6
Subjectcopper zinc superoxide dismutase; monoclonal antibody; mutant protein; amyotrophic lateral sclerosis; cell free system; cellular, subcellular and molecular biological phenomena and functions; immunoreactivity; intermolecular transmission; mesenchyme cell; mutation; protein expression; protein folding; protein protein interaction; cell line; murinae
AbstractHuman wild-type superoxide dismutase-1 (wtSOD1) is known to coaggregate with mutant SOD1 in familial amyotrophic lateral sclerosis (FALS), in double transgenic models of FALS, and in cell culture systems, but the structural determinants of this process are unclear. Here we molecularly dissect the effects of intracellular and cell-free obligately misfolded SOD1 mutant proteins on natively structured wild-type SOD1. Expression of the enzymatically inactive, natural familial ALS SOD1 mutations G127X and G85R in human mesenchymal and neural cell lines induces misfolding of wild-type natively structured SOD1, as indicated by: acquisition of immunoreactivity with SOD1 misfolding-specific monoclonal antibodies; markedly enhanced protease sensitivity suggestive of structural loosening; and nonnative disulfide-linked oligomer and multimer formation. Expression of G127X and G85R in mouse cell lines did not induce misfolding of murine wtSOD1, and a species restriction element for human wtSOD1 conversion was mapped to a region of sequence divergence in loop II and β-strand 3 of the SOD1 β-barrel (residues 24-36), then further refined surprisingly to a single tryptophan residue at codon 32 (W32) in human SOD1. Time course experiments enabled by W32 restriction revealed that G127X and misfolded wtSOD1 can induce misfolding of cell-endogenous wtSOD1. Finally, aggregated recombinant G127X is capable of inducing misfolding and protease sensitivity of recombinant human wtSOD1 in a cell-free system containing reducing and chelating agents; cell-free wtSOD1 conversion was also restricted by W32. These observations demonstrate that misfolded SOD1 can induce misfolding of natively structured wtSOD1 in a physiological intracellular milieu, consistent with a direct protein-protein interaction.
Publication date
LanguageEnglish
AffiliationNational Research Council Canada (NRC-CNRC); National Institute for Nanotechnology
Peer reviewedYes
NPARC number21271973
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Record identifierb6cedea5-4846-468f-8c10-dbab4d5fe78e
Record created2014-05-14
Record modified2016-05-09
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