Covalent binding of peroxidized phospholipid to protein: III. Reaction of individual phospholipids with different proteins

Various peroxidized phospholipids were reacted with proteins under N₂. In all cases, phospholipid is bound covalently to the proteins whose molecular size is increased. Both the amount of bound phospholipid and the increase in molecular size of the protein depends on the nature of the phospholipid. Ultraviolet (UV) absorption of the proteins is increased in qualitatively similar ways. Their difference spectra, which show a gradual increase in absorption from 400 nm toward shorter wavelength, differ from that of malonaldehyde-protein complexes. The various complexes of proteins and peroxidized phospholipids have similar fluorescence spectra showing two excitation maxima at 310–320 nm and at 340–350 nm, respectively, and emission maximum at ca. 400 nm. This is different from both fluorescence spectra of malonaldehyde-protein complexes and fluorescence spectra reported for proteins after reaction with peroxidized polyunsaturated fatty acids. Amino groups of the proteins are consumed in the reaction with peroxidized phospholipids. Blocking the amino groups decreases the binding of phospholipid considerably. Besides amino groups, other structures of the protein molecule react with the peroxidized phospholipids. The similar features of UV absorption, fluorescence, decrease of amino groups, and covalently bound phospholipid phosphorus of the various complexes suggest that they are formed by common type of reactions. The reactions seem to be different from those generally believed important between peroxidized lipid and protein. Important reacting species are compounds other than malonaldehyde. Preliminary report of this work was presented at the ISF/AOCS World Congress, New York, 1980.