The neurotoxicity of Aβ is associated with the formation of free radical by interacting with redox active metals such as Cu(2+). However, the relationship between ion-interaction, ion-driven free radical formation, and Aβ conformation remains to be further elucidated. In the present study, we investigated the correlation of Cu(2+) interaction and Cu(2+)-driven free radical formation with Aβ40 conformation. The Cu(2+)-binding affinity for Aβ40 in random coiled form is 3-fold higher than that in stable helical form. Unexpectedly but interestingly, we demonstrate in the first time that the stable helical form of Aβ40 can induce the formation of H(2)O(2) by interacting with Cu(2+). On the other hand, the H(2)O(2) generation is repressed at Aβ/Cu(2+) molar ratio ≥1 when Aβ40 adopts random coiled structure. Taken together, our result demonstrates that Aβ40 adopted a helical structure that may play a key factor for the formation of free radical with Cu(2+) ions.