期刊
JOURNAL OF MOLECULAR BIOLOGY
卷 385, 期 1, 页码 278-298出版社
ACADEMIC PRESS LTD- ELSEVIER SCIENCE LTD
DOI: 10.1016/j.jmb.2008.10.003
关键词
superoxide dismutase; amyotrophic lateral sclerosis; protein folding; metal-binding; dimer
资金
- Neuromuscular Research Partnership
- ALS Society of Canada, MDC and CIHR
More than 1.1.0 mutations in dimeric, Cu,Zn superoxide dismutase (SOD) have been linked to the fatal neurodegenerative disease, amyotrophic lateral sclerosis (ALS). In both human patients and mouse model studies, protein misfolding has been implicated in disease pathogenesis. A central step in understanding the misfolding/aggregation mechanism of this protein is the elucidation of the folding pathway of SOD. Here we report a systematic analyses of unfolding and folding kinetics using single- and double-jump experiments as well as measurements as a function of guanidium chloride, protein, and metal concentration for fully metallated (holo) pseudo wildtype and ALS-associated mutant (E100G,, G93R, G93A, and metal binding mutants G85R and H46R) SODs. The kinetic mechanism for holo SODs involves native dimer, monomer intermediate, and unfolded monomer, with variable metal dissociation from the monomeric states depending on solution conditions. The effects of the ALS mutations on the kinetics of the holoproteins in guanidium chloride are markedly different from those observed previously for acid-induced unfolding and for the unmetallated (apo) forms of the proteins. The mutations decrease the stability of holo SOD mainly by increasing unfolding rates, which is particularly pronounced for the metal-binding mutants, and have relatively smaller effects on the observed folding kinetics. Mutations also seem to favour increased formation of a Zn-free monomer intermediate, which has been implicated in the formation of toxic aggregates. The results reveal the kinetic basis for the extremely high stability of wild-type holo SOD and the possible consequences of kinetic changes for disease. (C) 2008 Elsevier Ltd. All rights reserved.
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