αB-Crystallin inhibits the cell toxicity associated with amyloid fibril formation by κ-casein and the amyloid-β peptide

Amyloid fibril formation is associated with diseases such as Alzheimer's, Parkinson's, and prion diseases. Inhibition of amyloid fibril formation by molecular chaperone proteins, such as the small heat-shock protein alpha B-crystallin, may play a protective role in preventing the toxicity associated with this form of protein misfolding. Reduced and carboxymethylated kappa-casein (RCM kappa-CN), a protein derived from milk, readily and reproducibly forms fibrils at physiological temperature and pH. We investigated the toxicity of fibril formation by RCM kappa-CN using neuronal model PC12 cells and determined whether the inhibition of fibril formation altered its cell toxicity. To resolve ambiguities in the literature, we also investigated whether fibril formation by amyloid-beta 1-40 (A beta(1-40)), the peptide associated with Alzheimer's disease, was inhibited by alpha B-crystallin and if this affected the toxicity of A beta. To this end, either RCM kappa-CN or A beta(1-40) was incubated at neutral pH to induce fibril formation before treating PC12 cells and assessing cell viability. Incubated (fibrillar) RCM kappa-CN was more toxic to PC12 cells than native RCM kappa-CN with the highest level of toxicity being associated with mature fibrils and protofibrils. Furthermore, the toxicity of RCM kappa-CN was attenuated when its fibril formation was inhibited, either through the chaperone action of alpha B-crystallin or when it interacted with its natural binding partners in milk, alpha(S)- and beta-casein. Likewise, incubating A beta(1-40) with alpha B-crystallin inhibited both A beta(1-40) fibril formation and the associated cell toxicity. Importantly, by inhibiting fibril formation, alpha B-crystallin prevents the cell toxicity associated with protein misfolding.