Mechanisms for Radiation Damage to DNA: LET Effects

描述（由申请人提供）：本研究的目的是阐明不同线性能量转移（LET）辐射对DNA造成辐射损伤的基本机制。我们的DNA辐射损伤的综合模型，描述了从最初形成的DNA离子自由基和激发态的事件，空穴和电子转移，糖自由基的形成，最后到分子产物将在每一步进行测试，以澄清导致DNA辐射损伤的基本过程。这些研究是在强调辐射直接影响的条件下进行的，将采用磁共振光谱、密度泛函理论和产物分析技术以及伽马和回旋加速器重离子束辐照。有三个目标：第一个目标将解决几个主要的未回答的问题，在DNA辐射损伤引起的洞。这一目标将专门采用C-8氘标记的定义序列寡核苷酸，以利用我们实验室最近的突破，使我们能够区分C-8氘标记的嘌呤碱基（鸟嘌呤或腺嘌呤）与未标记位点的空穴（阳离子自由基）。我们还发现，C-8标记允许鸟嘌呤和腺嘌呤阳离子自由基从它们的去质子化形式的区别。随着这些发展，我们会发现：A。空穴定位的碱基序列依赖性，B.在dsDNA中特定位点处鸟嘌呤和腺嘌呤阳离子自由基的质子化状态，c.空穴激发时碱基-碱基与碱基-糖转移的程度。我们的第二个目标将确定自由基形成和跟踪结构的LET在离子束照射的DNA的功能。我们将通过ESR光谱识别自由基，并确定它们的空间分布和聚类作为辐射沿着辐射轨迹的LET的函数。特别重要的是，将是最近发现的提示链断裂自由基，从糖磷酸骨架裂解的LET依赖性的研究。轨道核心中的自由基形成和聚集的性质与理解DNA中最重要的损伤（不可修复的多重损伤部位）的形成有关。我们的最终目标将采用理论计算，以进一步测试和确认在上述研究中提出的分子机制。特别重要的是将通过基离子自由基的激发态的TD-DFT理论进行处理，所述基离子自由基现在与DNA链断裂有关，并且随着辐射的LET增加而变得更加重要。我们相信，这一努力将使我们能够建立新的见解基本辐射过程的重要生物医学研究。 公共卫生相关性：本研究的目的是通过阐明不同线性能量转移（LET）辐射对DNA损伤的基本机制，建立一个全面的DNA辐射损伤模型。我们的DNA辐射损伤模型描述了从最初形成的DNA离子自由基和激发态，空穴和电子转移，糖基的形成，最后到分子产物的事件将在每一步进行测试，照亮导致DNA辐射损伤的基本过程。这些研究是在强调辐射的直接影响的条件下进行的，将采用伽马和回旋重离子束辐照、磁共振光谱、密度泛函理论和产物分析技术，并将解决对生物医学研究很重要的DNA辐射损伤方面尚未解决的主要问题。

DESCRIPTION (provided by applicant): The goal of this research is to elucidate fundamental mechanisms of radiation damage to DNA by radiations of varying linear energy transfer (LET). Our comprehensive model for DNA radiation damage that describes events from the initial formation of DNA ion radicals and excited states, to hole and electron transfer, to sugar radical formation and finally to molecular products will be tested at each step to clarify the fundamental processes resulting in DNA radiation damage. These studies, which are performed under conditions that emphasize the direct effect of radiation, will employ magnetic resonance spectroscopies, density functional theory and product analysis techniques as well as gamma and cyclotron heavy ion beam irradiations. There are three aims: The first aim will address several of the major unanswered questions in DNA radiation damage induced by holes. This aim will employ specifically C-8 deuterium labeled defined sequence oligos to exploit a recent breakthrough in our laboratory that allows us to distinguish a hole (cation radical) at a C-8 deuterium labeled purine base (guanine or adenine) from an unlabeled site. We have also found that the C-8 labeling allows the distinction of the guanine and adenine cation radicals from their deprotonated forms. With these developments we will find: a. the base sequence dependence of hole localization, b. the protonation states of guanine and adenine cation radicals at specific sites in dsDNA, c. the extent of base-to- base versus base-to-sugar transfer on hole excitation. Our second aim will identify radicals formed and track structure as a function of LET in ion beam irradiated DNA. We will identify radicals via ESR spectroscopy and ascertain their spatial distribution and clustering as a function of the LET of the radiation along the radiation track. Especially important will be a study of the LET dependence of recently discovered prompt strand break radicals that result from cleavage of the sugar phosphate backbone. The nature of the radical formation and clustering in the track core is pertinent to understanding the formation of the most important lesion in DNA the unrepairable multiply damaged site. Our final aim will employ theoretical calculations to further test and confirm molecular mechanisms proposed in the above studies. Especially significant will be treatment by TD- DFT theory of excited states of base ion radicals which are now implicated in DNA strand breaks and become more significant as the LET of the radiation increases. We believe this effort will allow us to establish new insights into fundamental radiation processes important for biomedical research. PUBLIC HEALTH RELEVANCE: The goal of this research is to develop a comprehensive model of DNA radiation damage by elucidating fundamental mechanisms of damage to DNA by radiations of varying linear energy transfer (LET). Our model for DNA radiation damage that describes events from the initial formation of DNA ion radicals and excited states, to hole and electron transfer, to sugar radical formation and finally to molecular products will be tested at each step to illuminate the fundamental processes resulting in DNA radiation damage. These studies, which are performed under conditions that emphasize the direct effect of radiation, will employ gamma and cyclotron heavy ion beam irradiations, magnetic resonance spectroscopies, density functional theory and product analysis techniques and will address major unanswered questions in DNA radiation damage important to biomedical research.