In the present study, we have investigated the proteome changes associated with glutamate‐induced HT22 cell death, a model system to study oxidative stress‐mediated toxicity. Among a number of HT22 proteins exhibiting altered expression, several molecular chaperones demonstrated substantial changes. For example, the levels of Hsp90 and Hsp70 decreased as cell death progressed whereas that of Hsp60 increased dramatically. Interestingly, cytosolic Hsp60 increased more prominently than mitochondrial Hsp60. Concomitantly, the accumulation of poly‐ubiquitylated proteins and differential regulation of the peptidase activities and the subunits of 26S proteasomes were observed in glutamate‐treated HT22 cells. Our findings that the molecular chaperones and the ubiquitin‐proteasome system undergo changes during glutamate‐induced HT22 cell death may suggest the importance of a protein quality control system in oxidative damage‐mediated toxicity.