Toxicology of Peptide/Protein Adduction by 4-Oxononenal

描述（由申请人提供）：本研究计划的目的是确定4-氧壬烯醛（4ONE），脂质过氧化的主要产物，对肽/蛋白质的反应性。4ONE在结构上类似于已知的细胞毒性脂质醛4-羟基壬烯醛（4HNE），但由于其化学结构，其活性可能更强。4HNE对蛋白质的修饰已被确定，并被认为是人类疾病（如动脉粥样硬化）发病机制的一个启动因素；然而，4ONE对肽/蛋白质的内聚尚未被研究甚至证实。为了实现本研究计划的目标，将测试以下工作假设：4ONE共价修饰具有残基特异性的肽和蛋白质，并可以抑制代谢4HNE的酶；此外，脂质过氧化在线粒体中产生的4ONE可以修饰在呼吸和代谢中重要的线粒体蛋白/酶。为了验证这一假设，将进行实验来鉴定4ONE加合物的肽残基，表征该化合物对特定氨基酸的偏好，并确定4ONE加合物的化学结构。将进行工作以确定4ONE是否抑制对4HNE代谢重要的酶。此外，将进行实验以确定通过线粒体脂质过氧化产生的4ONE是否会加合呼吸和代谢中重要的线粒体酶/蛋白质。这些目标的完成将为实现研究计划的目标提供科学数据，并评估4ONE蛋白内聚的毒理学/生理学意义。

DESCRIPTION (provided by applicant): The objective of this research proposal is to determine the reactivity of 4-oxononenal (4ONE), a major product of lipid peroxidation, toward peptides/protein. 4ONE is structurally analogous to the known cytotoxic lipid aldehyde 4-hydroxynonenal (4HNE) but is likely more reactive because of its chemical structure. Protein modification by 4HNE has been characterized and is thought to be an initiating factor in the pathogenesis of human diseases (e.g. atherosclerosis); however, the adduction of peptides/proteins by 4ONE has not been studied or even demonstrated. In order to fulfill the objective of this research proposal, the following working hypotheses will be tested: 4ONE covalently modifies peptides and proteins with residue specificity and can inhibit enzymes that metabolize 4HNE; furthermore, 4ONE generated in the mitochondria from lipid peroxidation can modify mitochonidrial proteins/enzymes important in respiration and metabolism. To test the hypothesis, experiments will be performed to identify peptide residues adducted by 4ONE, characterize the preference of the compound toward specific amino acids, and determine the chemical structure of 4ONE adducts. Work will be done to ascertain whether 4ONE inhibits enzymes important for metabolism of 4HNE. Furthermore, experiments will be conducted to determine whether 4ONE generated via mitochondrial lipid peroxidation adducts mitochondrial enzymes/proteins important in respiration and metabolism. Completion of these aims will provide scientific data to achieve the objective of the research proposal and assess the toxicological/physiological significance of protein adduction by 4ONE.