The Formation of Oxidatively Induced High-Molecular-Weight Aggregate of α-/γ-Crystallins

Abstract

α-/γ-Crystallin interactions under oxidation with ascorbate–FeCl3–EDTA–H2O2 followed by dialysis have been studied. A high-molecular-weight aggregate (HMWA) composed of α- and γ-crystallin was observed for the mixture of the dialyzed α-crystallin and the oxidized γ-crystallin through gel-filtration chromatography. This illustrates an interaction between α-crystallin and partially denatured γ-crystallin induced by oxidation. No HMWA formation was observed under the condition without dialysis and/or with the addition of catalase to the oxidized γ-crystallin prior to the addition of α-crystallin. More HMWA was formed by oxidized γ-crystallin followed by the addition of α-crystallin than by simultaneous oxidation of both α- and γ-crystallins. Conformational changes of α-crystallin during oxidation analyzed by circular dichroism spectra showed that oxidized α-crystallin can gradually be restored to an ordered structure through dialysis. The overall results imply that structural changes of both α- and γ-crystallins and dialysis are required to form HMWA. The observation of this oxidatively induced chaperone/substrate complex suggests that an efficient chaperone-like protective action against oxidative insults may exist in vivo.