Investigating the mechanism of ferritin protection of DNA

描述(申请人提供)：虽然衰老是一个复杂的、多维的过程，还没有完全了解，但DNA损伤的积累长期以来一直被认为是衰老的一个重要因素。铁储存蛋白铁蛋白在细胞的防御策略中发挥着重要作用。这些蛋白质会消耗亚铁和过氧化氢，否则会通过芬顿反应产生破坏性的氧自由基。虽然传统上认为铁蛋白只存在于细胞质中，但最近发现它存在于细胞核中，这为铁蛋白直接保护DNA开辟了新的可能性。本研究旨在阐明铁蛋白对DNA的保护作用机制。DPS是一种细菌铁蛋白，其DNA结合已被广泛研究，将被用作铁蛋白家族的模型。DNA通过其碱基进行电荷传递的能力已经被很好地研究，并且DNA电荷转移(CT)过程被认为在许多系统中具有生物学意义，例如氧化还原敏感转录因子的激活。在这项工作中，我们将确定铁蛋白DPS是否可以通过利用通过DNA的电荷传输来远距离保护基因组。这将通过将各种光谱与闪急技术相结合来实现，闪急技术是一种在现场产生一种能够氧化DNA的强大氧化剂的方法。在氧化时，缺乏电子(空穴)集中在鸟嘌呤上，鸟嘌呤是最容易被氧化的碱。然而，我们假设DNA结合的DPS可以通过DNA CT被氧化，以填补鸟嘌呤上的空洞并恢复DNA的完整性。瞬时吸收和电子顺磁共振光谱可以监测鸟嘌呤自由基和蛋白质氧化产物，因此可以用来确定铁蛋白是否可以通过在DNA介导的过程中被氧化来保护DNA。这项工作试图扩大一般的保护机制，包括远距离保护，希望阐明DNA损伤和修复之间动态相互作用的一个方面，这是导致衰老和与年龄相关的疾病的原因。

DESCRIPTION (provided by applicant): While aging is a complex, multidimensional process that is not yet fully understood, the accumulation of DNA damage has long been recognized as an important factor in aging. The iron storage protein ferritin is a significant player in the defensive strategy of the cell. These proteins deplete ferrous iron and hydrogen peroxide, which can otherwise produce damaging oxygen radicals via the Fenton reaction. While ferritin was traditionally believed to be present solely in the cytoplasm, it recently has been found in cell nuclei, opening up new possibilities for direct DNA protection by ferritin. This study will focus on elucidating the mechanism of ferritin protection of DNA. Dps, a bacterial ferritin whose DNA binding has been extensively studied, will be used as a model for the ferritin family. The ability of DNA to conduct charge through its base stack has been well-studied and DNA charge transfer (CT) processes have been proposed to be biologically relevant in a number of systems such as the activation of redox sensitive transcription factors. In this work, we will determine if the ferritin Dps can protect the genome from a distance by utilizing charge transport through DNA. This will be achieved by combining various spectroscopies with the flash-quench technique, a method to generate a powerful oxidant in situ that is capable of oxidizing DNA. Upon oxidation, the lack of an electron (hole) localizes on guanine, the most easily oxidized base. However, we hypothesize that DNA- bound Dps can become oxidized via DNA CT to fill the hole on the guanine and restore the integrity of the DNA. Transient absorption and electron paramagnetic resonance spectroscopies can monitor guanine radicals and protein oxidation products, and can therefore be used to determine if ferritin can protect DNA from a distance by becoming oxidized in a DNA- mediated process. This work seeks to expand the general mechanisms of protection to include protection from a distance, in the hopes of elucidating one aspect of the dynamic interplay between DNA damage and repair that contributes to aging and age-related disease.