A comprehensive study on the generation of reactive oxygen species in Cu-Aβ-catalyzed redox processes

In the amyloid plaques, a signature of AD, abnormally high Cu²⁺ concentrations are found bound to Aβ. Most of previous studies reported that Cu-Aβ could contribute to oxidative stress, as H₂O₂ and •OH are catalytically generated by Cu-Aβ with the assistance of biological reductant, with only one recent report stated that free O₂^•- is also generated in the Cu-Aβ catalyzed processes, where an indirect technique was applied. To comprehensively investigate the free radicals produced during this Cu-Aβ-mediated process with a biological reductant, DNA-cleavage assay, an indirect method, and two direct methods including electron paramagnetic resonance (EPR) spectroscopy and transient absorption spectroscopy (TAS), both having qualitative and quantitative power, were employed in this work. All the experimental results obtained from the three methods demonstrated that Cu-Aβ in the biological reducing environment was not only able to catalyze the production of H₂O₂ and •OH, but also to generate free O₂^•-. The results further indicated that O₂^•- was the precursor of H₂O₂ and •OH. It is also important to note that the results obtained from EPR spectroscopy and TAS provided direct evidence for the presence of O₂^•- and •OH. By virtue of the direct techniques, we also found that the longest peptide fragments of Aβ₁₆, Aβ₄₀, and Aβ₄₂ produced the least radicals with a lowest rate. More interestingly, the fibrillar forms of Aβ generated less O₂^•- and •OH compared with oligomeric and monomeric forms.