Structure/Function of Mn and Fe Superoxide Dismutases

说明(申请人提供)：超氧化物歧化酶(SOD)是含有锰、铁、铜/锌或镍活性中心的金属酶，用于保护生物系统免受有氧代谢产物超氧阴离子(O2)介导的氧化损伤。超氧化物歧化酶还被证明可以预防炎症，并参与一系列抗癌和抗衰老机制。这项建议侧重于结构上相似的依赖于锰和铁的超氧化物歧化酶，它们通过O2歧化成02和H202来完成其功能。在这项建议中概述的研究的长期目标是-确定对依赖于锰和铁的超氧化物歧化酶的反应活性的关键几何和电子结构贡献，以及-获得对这些酶的反应机理的分子水平的洞察。已经制定了以下具体目标：-生成被氧化和还原的Mn-和Fe-SOD活性中心的电子结构描述。-探索导致SOD金属专一性的因素。-评估第二球氨基酸在调节活性中心属性中的作用。-确定Mn-和Fe-SOD的底物-金属相互作用的性质，并在实验和理论水平上评估相应催化循环中的关键步骤。我们的方法涉及使用光谱工具(吸收、CD、MCD、EPR和RR)和计算方法(DFT和NBO)的组合来研究天然的Mn-和Fe-SOD蛋白质，催化失活的金属取代的锰和铁的超氧化物歧化酶物种，以及几个突变蛋白。这些研究将系统地探索影响超氧化物歧化酶活性的几何和电子因素。由于超氧化物歧化酶在疾病状态的动物模型中显示出治疗效果，低分子量超氧化物歧化酶模拟物(合成酶)已被提出用于治疗各种疾病。与天然的SOD酶相比，Synzyme作为药物制剂具有明显的优势，如细胞通透性、缺乏免疫原性、更长的寿命、潜在的口服给药和更低的生产成本。因此，了解Mn-和Fe-SOD酶实现其极高催化效率的原理，特别是第二配位壳氨基酸对活性中心电子学的影响，将有助于合理设计用于药物应用的SOD模拟物。

DESCRIPTION (provided by applicant): Superoxide dismutases (SODs) are metalloenzymes containing Mn, Fe, Cu/Zn, or Ni active sites that defend biological systems against oxidative damage mediated by the superoxide radical anion (O2), a product of aerobic metabolism. SODs have also been shown to protect against inflammation and are involved in a range of anti-cancer and anti-aging mechanisms. This proposal focuses on the structurally similar Mn- and Fe-dependent SODs, which accomplish their function through disproportionation of O2 to 02 and H202.The long-term objectives of the research outlined in this proposal are- to identify key geometric and electronic structure contributions to the reactivities of Mn- and Fe-dependent SODs and- to obtain molecular-level insight into the reaction mechanisms of these enzymes.With these goals in mind, the following specific aims have been formulated:- Generate electronic structure descriptions of the oxidized and reduced Mn- and Fe-SOD active sites.- Explore the factors responsible for SOD metal specificity.- Assess the role of second-sphere amino acids in tuning active site properties.- Define the nature of the substrate-metal interaction for Mn- and Fe-SODs and evaluate key steps in the corresponding catalytic cycles on experimental and theoretical levels.Our approach involves using a combination of spectroscopic tools (absorption, CD, MCD, EPR, and rR) and computational methods (DFT and NBO) to study the native Mn- and Fe-SOD proteins, the catalytically inactive metal-substituted Mn- and Fe-SOD species, and several mutant proteins. These studies will systematically explore geometric and electronic factors contributing to SOD activity.As SOD enzymes demonstrate therapeutic efficacy in animal models of disease states involving superoxide, low molecular weight SOD enzyme mimics (synzymes) have been proposed for the treatment of a variety of diseases. Synzymes could have distinct advantages over natural SOD enzymes as pharmaceutical agents, such as cellular permeability, lack of immunogenicity, longer lifetimes, potential for oral delivery, and lower production costs. Thus, understanding the principles by which the Mn- and Fe-SOD enzymes achieve their remarkably high catalytic rates, in particular the influence of second coordination shell amino acids on active site electronics, could aid significantly in the rational design of SOD mimics for pharmaceutical applications.