Incubation of glucose-6-phosphate dehydrogenase (Glu-6-PDH) from Leuconostoc mesenteroides with the lipid peroxidation product 4-hydroxy-2-nonenal leads to the formation of cross-linked protein. This is accompanied by the appearance of protein-associated fluorescence with excitation and emission maxima of 340 and 415 nm, respectively, and with the disappearance of histidine and lysine residues. Cross-linked protein is less susceptible than native Glu-6-PDH to proteolysis by the multicatalytic protease, a multienzymic proteolytic complex involved in the intracellular degradation of damaged proteins. In addition, 4-hydroxy-2-nonenal-modified Glu-6-PDH inhibits the multicatalytic protease and can therefore prevent the efficient degradation of oxidized protein. These findings may have important implications for the accumulation of altered protein and fluorescent material in vivo, processes that are believed to be involved in age- and disease-related impairment of cellular function.