INACTIVATION OF RIBONUCLEASE AND OTHER ENZYMES BY PEROXIDIZING LIPIDS AND BY MALONALDEHYDE

Quantitative enzyme inactivation by lipid peroxidation has been studied. Sulfhydryl enzymes are most susceptible to inactivation by lipid peroxidation intermediates. Oxidation products of polyunsaturated lipids also inactivate nonsulfhydryl enzymes, for example, ribonuclease A. Concomitant with the loss of ribonuclease A activity is the appearance of fluorescence in the enzyme-lipid reaction mixture. The inactivated RNase A shows fluorescent monomer, dimer, and higher molecular weight species in the Sephadex G-100 fractionation pattern. The fluorescence maximum is at 470 mμ; and the excitation maximum is at 395 mg. Ribonuclease A, inactivated by malonaldehyde, has fluorescence and a gel filtration pattern similar to that of the enzyme inactivated by peroxidizing polyunsaturated lipids. Malonaldehyde is probably the agent responsible for the intra- and intermolecular cross-linking of ribonuclease A. The fluorescence produced from the cross-linking is attributed to the conjugated imine structure formed in protein between two ϵ-amino groups and malonaldehyde. There are marked similarities between the ribonuclease A-polyunsaturated lipid product and age pigment; cardiac age pigment is a protein-lipid complex and both have similar fluorescence characteristics. © 1969, American Chemical Society. All rights reserved.