REACTIONS OF 4-HYDROXY-2(E)-NONENAL AND RELATED ALDEHYDES WITH PROTEINS STUDIED BY C-13 NUCLEAR-MAGNETIC-RESONANCE SPECTROSCOPY

In order to understand the modifications of proteins produced by aldeh ydes of lipid peroxidation, [1-C-13]-2(E)-hexenal, [1-C-13]-4-oxopenta nal, and a mixture of [1-C-13]- and [2-C-13]-4-hydroxynon-2(E)-enal we re synthesized and the reaction of each of the labeled aldehydes with bovine serum albumin was analyzed by C-13 NMR spectroscopy. Protein nu cleophiles add to the 3-position of hexenal, and the resulting propana l moieties appear to undergo aldol condensation, form imine cross-link s with lysyl residues, or lead to pyridinium rings. During the reactio n of 4-oxopentanal with the lysyl residues of bovine serum albumin, on ly 1-alkyl-2-methylpyrrole and a possible intermediate leading to the pyrrole were observed. Hydroxypyrrolidine cross-links such as 25 could not be detected, leaving the pyrrole as the mediator of protein cross -linking. The Michael adducts are the major products in the reaction b etween 4-hydroxynon-2-enal and proteins. They exist almost exclusively in the cyclic hemiacetal form and do not appear to cross-link through imine formation with lysyl residues. A minor pathway involves the rea ction of 4-hydroxynon-2-enal with the lysyl amino groups of protein re sulting in 2-pentylpyrrole adducts that may mediate protein cross-link ing. The Michael adducts appear not to be the direct source of the pyr role, but the imine 32 and the enamine 35 are likely intermediates tow ard the five-membered ring.