4-Hydroperoxy-2-nonenal Is Not Just an Intermediate but a Reactive Molecule That Covalently Modifies Proteins to Generate Unique Intramolecular Oxidation Products

Graduate School of Bioagricultural Sciences, Nagoya University, Nagoya 464-8601, Japan.
Journal of Biological Chemistry (Impact Factor: 4.57). 06/2011; 286(33):29313-24. DOI: 10.1074/jbc.M111.255737
Source: PubMed


α,β-Unsaturated aldehydes generated during lipid peroxidation, such as 4-oxoalkenals and 4-hydroxyalkenals, can give rise to protein degeneration in a variety of pathological states. Although the covalent modification of proteins by these end products has been well studied, the reactivity of unstable intermediates possessing a hydroperoxy group, such as 4-hydroperoxy-2-nonenal (HPNE), with protein has received little attention. We have now established a unique protein modification in which the 4-hydroperoxy group of HPNE is involved in the formation of structurally unusual lysine adducts. In addition, we showed that one of the HPNE-specific lysine adducts constitutes the epitope of a monoclonal antibody raised against the HPNE-modified protein. Upon incubation with bovine serum albumin, HPNE preferentially reacted with the lysine residues. By employing N(α)-benzoylglycyl-lysine, we detected two major products containing one HPNE molecule per peptide. Based on the chemical and spectroscopic evidence, the products were identified to be the N(α)-benzoylglycyl derivatives of N(ε)-4-hydroxynonanoic acid-lysine and N(ε)-4-hydroxy-(2Z)-nonenoyllysine, both of which are suggested to be formed through mechanisms in which the initial HPNE-lysine adducts undergo Baeyer-Villiger-like reactions proceeding through an intramolecular oxidation catalyzed by the hydroperoxy group. On the other hand, using an HPNE-modified protein as the immunogen, we raised a monoclonal antibody against the HPNE-modified protein and identified one of the HPNE-specific lysine adducts, N(ε)-4-hydroxynonanoic acid-lysine, as an intrinsic epitope of the monoclonal antibody. Furthermore, we demonstrated that the HPNE-specific epitopes were produced not only in the oxidized low density lipoprotein in vitro but also in the atherosclerotic lesions. These results indicated that HPNE is not just an intermediate but also a reactive molecule that could covalently modify proteins in biological systems.

4 Reads
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Oxygen is essential for the growth and function of mammalian cells. However, imbalances in oxygen or abnormalities in the ability of a cell to respond to oxygen levels can result in oxidative stress. Oxidative stress plays an important role in a number of diseases including atherosclerosis, rheumatoid arthritis, cancer, neurodegenerative diseases and asthma. When membrane lipids are exposed to high levels of oxygen or derived oxidants, they undergo lipid peroxidation to generate oxidized phospholipids (oxPL). Continual exposure to oxidants and decomposition of oxPL results in the formation of reactive electrophiles, such as 4-hydroxy-2-nonenal (HNE). Reactive lipid electrophiles have been shown to covalently modify DNA and proteins. Furthermore, exposure of cells to lipid electrophiles results in the activation of cytoprotective signaling pathways in order to promote cell survival and recovery from oxidant stress. However, if not properly managed by cellular detoxification mechanisms, the continual exposure of cells to electrophiles results in cytotoxicity. The following perspective will discuss the biological importance of lipid electrophile protein adducts including current strategies employed to identify and isolate protein adducts of lipid electrophiles as well as approaches to define cellular signaling mechanisms altered upon exposure to electrophiles. This article is part of a Special Issue entitled: Oxidized phospholipids-their properties and interactions with proteins.
    Biochimica et Biophysica Acta 04/2012; 1818(10):2424-35. DOI:10.1016/j.bbamem.2012.04.014 · 4.66 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: 4-Hydroxynonenal (HNE) is the most abundant and toxic aldehyde generated by the oxidation of plasma membrane polyunsaturated fatty acids. Systemic lupus erythematosus (SLE), a chronic autoimmune disease, is primarily characterized by increased levels of autoantibodies, predominantly against ds-DNA. However, the initial antigenic stimulus for the disease etiopathogenesis has remained elusive. HNE has been extensively used as a biomarker of oxidative stress. It can form adduct with proteins, making them highly immunogenic. Increased levels of such aldehyde-protein adducts have been reported in various pathological states, including autoimmune disorders like SLE and arthritis. In the present study, HNE-mediated structural changes in human serum albumin (HSA) were characterized by UV, fluorescence, CD and FT-IR spectroscopy as well as by polyacrylamide gel electrophoresis. Furthermore, immunogenicity of native and HNE-modified HSA was probed in female rabbits. The HNE-modified HSA was highly immunogenic eliciting high titre immunogen specific antibodies. Binding of SLE anti-DNA antibodies was analyzed by direct binding and competition ELISA. The data show preferential binding of SLE autoantibodies to HNE-modified HSA as compared to native HSA or native DNA. Our results suggest that HNE modification generates neoepitopes on HSA causing enhanced autoantibodies production. The results point towards the possible role of HNE-modified HSA in SLE etiopathogenesis.
    Human immunology 08/2012; 73(11):1132-9. DOI:10.1016/j.humimm.2012.08.011 · 2.14 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Diabetes has emerged as a major threat to worldwide health. The exact mechanisms underlying the disease are unknown; however, there is growing evidence that the excess generation of reactive oxygen species (ROS) associated with hyperglycemia, causes oxidative stress in a variety of tissues. In this context, various natural compounds with pleiotropic actions like α-lipoic acid (LA) are of interest, especially in metabolic diseases such as diabetes. LA, either as a dietary supplement or a therapeutic agent, modulates redox potential because of its ability to match the redox status between different subcellular compartments as well as extracellularly. Both the oxidized (disulfide) and reduced (di-thiol: dihydro-lipoic acid, DHLA) forms of LA show antioxidant properties. LA exerts antioxidant effects in biological systems through ROS quenching but also via an action on transition metal chelation. Dietary supplementation with LA has been successfully employed in a variety of in vivo models of disease associated with an imbalance of redox status: diabetes and cardiovascular diseases. The complex and intimate association between increased oxidative stress and increased inflammation in related disorders such as diabetes, makes it difficult to establish the temporal sequence of the relationship.
    Molecular Nutrition & Food Research 01/2013; 57(1):114-25. DOI:10.1002/mnfr.201200608 · 4.60 Impact Factor
Show more

Similar Publications