( E)-4-Hydroxynon-2-enal (HNE), an electrophilic bifunctional cytotoxic lipid peroxidation product, forms covalent adducts with nucleophilic side chains of amino acid residues. HNE-derived adducts have been implicated in many pathophysiological processes including atherosclerosis, diabetes, and Alzheimer's disease. Tritium- and deuterium-labeled HNE ( d 4-HNE) were used orthogonally to study adduction with proteins and individual nucleophilic groups of histidyl, lysyl, and cysteine residues. Using tritium-labeled HNE, we detected the binding of 486 molecules of HNE per low-density lipoprotein (LDL) particle, significantly more than the total number of all reactive nucleophiles in the LDL particle. This suggests the formation of adducts that incorporate multiple molecules of HNE with some nucleophilic amino acid side chains. We also found that the reaction of a 1:1 mixture of d 4-HNE and d 0-HNE with N-acetylhistidine, N-acetyl-Gly-Lys-OMe, or N-acetyl cysteine generates 1:1, 2:1, and 3:1 adducts, which exhibit unique mass spectral signatures that aid in structural characterization. A domino-like reaction of initial 1:1 HNE Michael adducts of histidyl or lysyl nucleophiles with multiple additional HNE molecules forms 2:1 and 3:1 adducts that were structurally characterized by tandem mass spectrometry.

中文翻译：

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低密度脂蛋白具有结合 (E)-4-hydroxynon-2-enal (HNE) 的巨大能力：检测和表征含有多个 HNE 分子的赖氨酰和组氨酰加合物。

( E)-4-Hydroxynon-2-enal (HNE) 是一种亲电双功能细胞毒性脂质过氧化产物，与氨基酸残基的亲核侧链形成共价加合物。HNE 衍生的加合物与许多病理生理过程有关，包括动脉粥样硬化、糖尿病和阿尔茨海默病。氚和氘标记的 HNE (d 4-HNE) 被正交用于研究与蛋白质和组氨酰、赖氨酰和半胱氨酸残基的单个亲核基团的加成反应。使用氚标记的 HNE，我们检测到每个低密度脂蛋白 (LDL) 颗粒与 486 个 HNE 分子的结合，显着超过 LDL 颗粒中所有反应性亲核试剂的总数。这表明形成了包含多个 HNE 分子和一些亲核氨基酸侧链的加合物。我们还发现 d 4-HNE 和 d 0-HNE 的 1:1 混合物与 N-乙酰组氨酸、N-乙酰-Gly-Lys-OMe 或 N-乙酰半胱氨酸的反应生成 1:1、2:1和 3:1 加合物，它们表现出独特的质谱特征，有助于结构表征。组氨酰或赖氨酰亲核试剂的初始 1:1 HNE Michael 加合物与多个附加 HNE 分子的多米诺样反应形成 2:1 和 3:1 加合物，其结构通过串联质谱法表征。