Model Studies on Protein Side Chain Modification by 4-Oxo-2-nonenal

Abstract

trans-4-Oxo-2-nonenal (ONE) has recently been demonstrated to be a direct product of lipid peroxidation. In earlier studies to elucidate the structure of the trans-4-hydroxy-2-nonenal (HNE)-derived fluorescent Lys-Lys cross-link, we showed that ONE was capable of both oxidative and nonoxidative cross-linking of amines. A more comprehensive study on nonoxidative modification of protein nucleophiles by ONE is described here, focusing on the initial Michael addition of imidazole, thiol, and amine groups to C2 or C3 to give 4-keto aldehydes that can then condense with amines to form nucleophile-substituted pyrroles. 2,3-Substituted pyrroles (major) and 2,4-substituted pyrroles (minor) were distinguished by 2D NMR techniques, and N(tau)-substitution is preferred over N(pi)-substitution in the Michael addition of histidine. Mechanisms of both nonoxidative and oxidative side chain reactions of ONE are discussed, as is the relative propensity (ONE > HNE) to induce cross-linking of the model proteins ribonuclease A and beta-lactoglobulin.

Lipid Peroxidation Modification of Protein Generates Nϵ-(4-Oxononanoyl)lysine as a Pro-inflammatory Ligand

“…Using the Lys derivative, they also have determined the relative yield of the various ONE adducts as a function of time and found that the ONL adduct was shown to survive at the expense of the short lived Schiff base and Michael adducts. Although the ONL represents the major long lived noncross-linked ONE modification on the Lys residues (23), previous studies of the ONE that missed this ONL adduct either mainly focused on the early and/or reversibly formed ONEderived adducts (11) or on adducts that could be readily detected in preparative isolation work by characteristic UVvisible spectral and NMR spectrometric signatures (21,41,42). In addition to ONE, 9,12-dioxo-10(E)-dodecaenoic acid, an ONE-like lipid peroxidation product that contains the carboxyl terminus, covalently reacts with the Lys residues to generate the same 4-ketoamide-type adduct (23).…”

Oxidative Stress and Covalent Modification of Protein with Bioactive Aldehydes

“…Although 4-HNE has historically been the most well studied lipid peroxidation product (11), 4-ONE is also highly reactive (12). After forming Michael adducts, the aldehyde moiety may in some cases undergo Schiff base formation with amines of adjacent lysines, producing intra-and/or intermolecular cross-linked amino acids (13,14). In the case of 4-ONE, modification of lysine residues through 1,2-addition (Schiff base formation and the addition of water to the double bond) can also result in ketoamide adducts (15).…”

Reaction Products of γ-Tocopherol with (E)-4-Oxo-2-nonenal in Acidic Acetonitrile

“…18,23,[28][29][30][31] The addition by nucleobases also occurs at the C-1 position of ONE with the subsequent reaction at the C-2 position of the double bond to form 2-oxoheptyl-substituted etheno adducts. 19,[32][33][34][35][36] The acid-catalyzed reaction of -tocopherol with ONE in an acetonitrile solution proceeded by a different mechanism (Fig.…”

“…The first step would be isomerization of the double bond of ONE from trans to cis, this being followed by the formation of the enol tautomer, 4-hydroxy-2,4-nonadienal. 23) Intramolecular nucleophilic addition between the 4-hydroxyl and 1-carbonyl groups of the enol molecule would then give a cyclic hemiacetal intermediate. If -tocopherol is present in the reaction mixture, the C-5 position of -tocopherol could attack the double bond at the C-5 A reaction mixture of -tocopherol (10 mM) and 2-(1-hydroxypentyl)furan (20 mM) in acetonitrile was incubated at 37 C for 3 h in the absence (A) or presence (B) of 50 mM HCl.…”

“…Compound 4 has already been reported to be the major ONE-derived side product observed in reactions with some nucleophiles. 23) When the reaction proceeds or the reaction is performed under stronger acidic conditions, cleavage of the furan ring of compound 1 occurs and the attack by the 6-hydroxyl group of the -tocopherol moiety at the C-4 position of the opened furan ring results in the formation of compound 2.…”