Document Detail


Pro-Oxidant Copper-Binding Mode of the Apo Form of ALS-Linked SOD1 Mutant H43R Denatured at Physiological Temperature.
MedLine Citation:
PMID:  23837654     Owner:  NLM     Status:  Publisher    
Abstract/OtherAbstract:
The mutation of Cu,Zn-superoxide dismutase (SOD1), a major antioxidant enzyme, is associated with amyotrophic lateral sclerosis (ALS). In a previous study, we showed that the metal-depleted apo form of an ALS-linked mutant, H43R, undergoes denaturation at physiological temperature (37 °C) in 90 min and acquires pro-oxidant activity in the presence of Cu(2+) and H2O2. In this study, we have examined the Cu(2+)-binding mode of denatured apo-H43R by circular dichroism (CD), fluorescent oxidation, UV Raman spectroscopy, and photooxidation. CD spectroscopy indicates that denatured apo-H43R loses native β-barrel structure and the binding of Cu(2+) to the denatured apo form induces local refolding. Fluorescent-oxidation assays in the absence and presence of Cu(2+) chelators show that denatured apo-H43R contains two Cu(2+)-binding sites with higher and lower Cu(2+) affinities and with pro-oxidant activities in the reverse order. UV Raman spectroscopy gives evidence that His residues are bound to Cu(2+) mainly through the imidazole Nτ atom at the higher-affinity site and through the Nπ atom at the lower-affinity site, sharing one His residue with each other. The Cu(2+)-binding mode of denatured apo-H43R is analogous to but different from the Cu,Zn-binding mode of the native holo form. Photooxidation experiments confirm the involvement of His residues in the pro-oxidant activity. Taken together, it is suggested that the binding of Cu(2+) induces the local refolding of denatured apo-H43R to create toxic catalytic centers that convert the enzyme from antioxidant to pro-oxidant, leading to the pathogenesis of ALS. His residues are essential for both Cu(2+)-binding and pro-oxidant activities.
Authors:
Nobuhiro Fujimaki; Furi Kitamura; Hideo Takeuchi
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Publication Detail:
Type:  JOURNAL ARTICLE     Date:  2013-7-22
Journal Detail:
Title:  Biochemistry     Volume:  -     ISSN:  1520-4995     ISO Abbreviation:  Biochemistry     Publication Date:  2013 Jul 
Date Detail:
Created Date:  2013-7-23     Completed Date:  -     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  0370623     Medline TA:  Biochemistry     Country:  -    
Other Details:
Languages:  ENG     Pagination:  -     Citation Subset:  -    
Affiliation:
Graduate School of Pharmaceutical Sciences, Tohoku University , Aobayama, Sendai 980-8578, Japan.
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