Biological consequences of protein oxidation

过氧化氢（H2O2）等氧化剂参与介导多种人类疾病，包括动脉粥样硬化、癌症和神经退行性疾病。氧化剂通过破坏生物分子和改变细胞代谢来促进疾病进程。氧化损伤的关键靶点是结构蛋白和酶。为了了解氧化应激如何导致疾病，重要的是要发现哪些蛋白质受到氧化应激的影响，它们被修饰到什么程度，以及修饰的功能后果。先前关于蛋白质氧化的研究涉及将纯化蛋白质暴露于氧化剂来源（例如铁/抗坏血酸）并测量损害的程度和后果。最近，我们致力于研究暴露于化疗药物的肿瘤细胞中蛋白质氧化的诱导。文献报道表明，化疗药物诱导细胞凋亡是由细胞内产生的氧化剂介导的。我们着手确定蛋白质是否在这一过程中被氧化，如果是的话，特定蛋白质的氧化修饰是否有助于凋亡过程。在过去的一年里，研究人员进行了实验，以确定在化疗诱导的细胞凋亡和h2o2诱导的坏死过程中可能发生氧化的特定蛋白质。重点放在鉴定线粒体蛋白，可能会被修改，因为线粒体功能是如此核心的凋亡过程。药物VP-16诱导细胞凋亡，通过测定蛋白质羰基（Western blot assay）和总蛋氨酸亚砜来评估蛋白质氧化（与NIH NHLBI的研究人员合作）。到目前为止，我们还没有发现vp -16诱导的细胞凋亡过程中蛋白质氧化增加的证据。也没有诱导脂质过氧化，抗氧化化合物对抑制vp -16诱导的细胞死亡无效。我们的研究结果证明细胞凋亡并不一定涉及氧化应激，也不需要蛋白质氧化。这一发现意义重大，因为它挑战了目前认为细胞内氧化剂的形成是细胞凋亡过程不可分割的一部分的教条。这项工作目前正在准备发表在同行评议的期刊上。

Oxidants such as hydrogen peroxide (H2O2) are implicated in mediating a wide array of human diseases including atherosclerosis, cancer, and neurodegenerative diseases. Oxidants contribute to disease processes by causing damage to biomolecules and altering cellular metabolism. Key among the targets for oxidative damage are structural proteins and enzymes. In order to understand how oxidative stress can cause disease, it is important to discover which proteins become affected by oxidative stress, to what degree they are modified, and the functional consequences of the modifications. Previous studies on protein oxidation involved exposing purified proteins to a source of oxidants (e.g., iron/ascorbate) and measuring the extent and consequences of the damage. More recently, we have directed our efforts towards studying induction of protein oxidation in tumor cells exposed to chemotherapy drugs. Reports in the literature suggest that induction of apoptosis by chemotherapeutic agents is mediated by oxidants generated within the cells. We set out to determine whether proteins become oxidized in this process and, if so, whether oxidative modification of specific proteins contributes to the apoptotic process. Over the past year, experiments were carried out to identify specific proteins which might undergo oxidation during chemotherapy-induced apoptosis and H2O2-induced necrosis. Emphasis was placed on identifying mitochondrial proteins that might become modified since mitochondrial function is so central to the apoptotic process. Apoptosis was induced with the drug VP-16 and protein oxidation was assessed by measuring protein carbonyls (Western blot assay) and total methionine sulfoxide (in collaboration with investigators at the NHLBI, NIH). To date, we have found no evidence of increased protein oxidation during VP-16-induced apoptosis. There was also no induction of lipid peroxidation, and antioxidant compounds were ineffective at inhibiting VP-16-induced cell death. Our results provide evidence that apoptosis does not necessarily involve oxidative stress and does not require protein oxidation. This finding is significant because it challenges the current dogma which suggests that formation of intracellular oxidants is an integral part of the apoptotic process. The work is currently being prepared for publication in a peer-reviewed journal.