Mechanisms of electron transfer in nitric oxide synthases

一氧化氮合酶(NOS)，一种黄血球蛋白，严格调节一氧化氮(NO)的合成，从而 它作为血管扩张和神经传递的关键信号分子的双重生物学活性 在较高浓度下，也可作为防御性细胞毒素。三种一氧化氮合酶亚型，iNOS，eNOS 和nNOS(诱导型、内皮型和神经型)，通过严格的调控实现其关键的生物学功能 通过域间相互作用的域间电子转移(IET)过程。我们的长期目标是确定 一氧化氮合酶催化的分子机制，包括结构域间相互作用如何调节 催化相关的IET过程和一氧化氮合酶的功能。 在这个项目中，我们将重点研究FMN结构域与血红素对接的重要作用 结构域在调节一氧化氮合酶亚型功能中的作用。一氧化氮合酶的一个令人兴奋的最新发展是 发现结构域间FMN-血红素相互作用对调节FMN的反应性和结构的重要性 催化亚铁血红素活性中心。最近的发现提出了两个新的重要机制问题，必须 旨在了解一氧化氮合酶的调节机制：1)特定的域间相互作用控制和 促进FMN结构域与血红素结构域的有效对接？2)FMN结构域如何 对接，除了控制催化必需的FMN-血红素IET外，还调节血红素的反应性 活动站点？我们的两个协同和互补的目标旨在直接解决这些重要的问题 问题。我们将使用脉冲电子顺磁共振、激光闪光的集成程序 光解和突变研究。在缺乏全长哺乳动物一氧化氮合酶结构的情况下， 这种方法对于拟议的研究来说是非常合适的。 这个项目寻求解决与一氧化氮合酶相关的基本问题。我们的研究将确定 通过调节FMN和FMN的排列控制一氧化氮合酶异构体活性的关键结构决定因素 亚铁血红素区域。新的信息可以促进新的选择性激活剂的生物理性开发或 这些临床重要酶的抑制剂，以便提供更好的治疗干预措施。在……里面 按照R15计划的指导方针，我们将继续培养我们的21世纪生物医学人才 科学家通过这一多学科的项目。

Nitric oxide synthase (NOS), a flavo-hemoprotein, tightly regulates nitric oxide (NO) synthesis and thereby its dual biological activities as a key signaling molecule for vasodilatation and neurotransmission at low concentrations, and also as a defensive cytotoxin at higher concentrations. Three NOS isoforms, iNOS, eNOS and nNOS (inducible, endothelial, and neuronal NOS), achieve their key biological functions by tight regulation of interdomain electron transfer (IET) process via interdomain interactions. Our long-term goal is to determine the molecular mechanism of NOS enzyme catalysis, including how the interdomain interactions modulate the catalytically relevant IET processes and the NOS enzyme function. In this project we will focus on investigating important roles of the FMN domain docking to the heme domain in regulating the NOS isoform function. An exciting recent development in the NOS enzymes is the discovery of importance of the interdomain FMN-heme interactions for modulating reactivity and structure of the catalytic heme active site. The recent findings raise two new important mechanistic questions that must be addressed to understand NOS regulation mechanism: 1) what specific interdomain interactions govern and facilitate the productive docking of the FMN domain to the heme domain? 2) how can the FMN domain docking, in addition to controlling the catalytically essential FMN-heme IET, modulate reactivity of the heme active site? Our two synergistic and complementary Aims are designed to directly address these important questions. We will employ an integrated program of pulsed electron paramagnetic resonance, laser flash photolysis and mutational studies. In the absence of a structure of full-length mammalian NOS, the combined approach is highly appropriate for the proposed study. This project seeks to address fundamental questions relating to the NOS enzymes. Our study will identify key structural determinants in controlling NOS isoform activity by modulating the alignment of the FMN and heme domains. The new information could facilitate bio-rational development of new selective activators or inhibitors for these clinically important enzymes, in order to provide better therapeutic interventions. In accordance with the guideline of the R15 program we will continue to train our 21st century biomedical scientists by this multi-disciplinary project.