Arg199: key player in electron transfer in NOS enzyme

描述(申请人提供)：一氧化氮合酶家族催化L-精氨酸转化为L-瓜氨酸和一氧化氮(NO)，这是一种重要的细胞信使分子，与多种病理生理条件有关，如感染性休克、炎症性功能障碍、神经退行性疾病状态，以及最重要的心血管疾病。然而，尽管人们对NO在各种疾病中的作用有了越来越多的了解，但有效的治疗方法仍有待于完全了解NOS内的结构-功能关系。这一认识对于通过设计更有效的调节药物来调节体内内源性NO的产生是必不可少的。有人认为，一氧化氮合酶活性部位具有独特的协同结构功能和电子性质(动态和协同工作)，专为一氧化氮合酶的功能而调节。这个系统的复杂性要求使用复杂的计算方法，以充分了解和表征这种酶的作用机制。然而，到目前为止，利用第一原理量子力学(QM)和复合量子力学/分子力学(QM/MM)方法，结合分子动力学(MD)模拟和各种其他计算分析工具，进行全面的全面计算研究尚未得到利用。因此，这项提议的目的有三个方面：(1)通过MD模拟研究NOS环境，以全面表征四氢生物蝶呤(H4B)辅因子(精氨酸氧化的及时电子源)的动态行为，其自由基物种先前已在电子自旋共振实验中被确定；(2)通过QM和QM/MM计算和电子转移途径模拟，分析促进该自由基物种形成的酶环境，以确定腔内促进辅因子向血红素亚铁-二氧基中间体的电子转移的关键分子作用者；(3)通过计算来研究和阐明，这些关键角色在一氧化氮合酶的电子转移中的作用，试图指导未来的实验和药物干预，以抑制这一重要的酶。主要的焦点放在Arg199如何调节一氧化氮合酶中的电子转移上，这一作用只在系统的动力学过程中才被揭示。

DESCRIPTION (provided by applicant): The family of nitric oxide synthases (NOS) catalyzes the conversion of L-arginine to L-citrulline and nitric oxide (NO), an important cellular messenger molecule, which has been implicated in a variety of pathophysiological conditions such as septic shock, inflammatory dysfunction, neurodegenerative disease states, and, most importantly, cardiovascular disease. However, despite the increased understanding of the role of NO in contributing to various conditions, effective therapies still await the complete understanding of structure-function relationships within NOS. This understanding is imperative for the modulation of endogenous NO production in vivo, through the design of more effective regulatory drugs. It is proposed that the NOS active site possesses unique synergetic structural-function and electronic properties (which work together both dynamically and cooperatively) specifically tuned for the NOS enzyme functionality. The complexities of this system require the use of sophisticated computational methodologies, in order to fully understand and characterize the functioning mechanism of this enzyme. However, to date, a full-length comprehensive computational investigation, using first principle quantum mechanical (QM) and composite quantum mechanical and molecular mechanics (QM/MM) methodologies, coupled with molecular dynamics (MD) simulation and a variety of other computational analysis tools, has yet to be utilized. Therefore, the intent of this proposal is three fold: (1) to study the NOS environment through MD simulations, in order to fully characterize the dynamic behavior of the tetrahydrobiopterin (H4B) cofactor (a timely electron source for arginine oxidation), whose radical species has been previously identified in electron spin resonance experiments, (2) to analyze the enzyme environment that facilitates the formation of this radical species through QM and QM/MM calculations and electron transfer pathway modeling, in order to identify the key molecular players within the cavity which promote the electron transfer from the cofactor to the heme ferrous-dioxy intermediate, (3) to investigate and elucidate, through computation, the roles of these key players in the electron transfer within NOS, in attempts to direct future experimental and pharmacological interventions into the inhibition of this important enzyme. Primary focus is placed on how Arg199 modulates the electron transfer in NOS a role only revealed during the dynamics of the system.