Principles of C-H and O2 Activation

描述(由申请人提供)：本申请的主要目的是阐明支配酶催化的C-H和O2活化过程的原理。对C-H活化(质子、氢化物和氢原子)的研究主要集中在理解量子力学氢转移的作用及其与蛋白质结构和动力学的联系。氧活化的研究针对的是关于氧物种的性质和/或导致产物形成的反应途径仍然特别神秘的反应。具有重要药理意义的脂氧合酶已经成为旨在了解氢隧道在酶催化中的作用的研究的中心；未来的研究包括对蛋白质运动及其与隧道事件的联系的探测，高压对隧道事件的影响，以及与中温大豆脂肪氧合酶相比较的嗜热同系物的特征。与底物结合口袋垂直的蛋白质离散区域内的氧通道已在脂氧合酶中被假定；这将通过定点突变进行进一步测试。酪胺单加氧酶(T2M)是真核生物铜蛋白家族中的一员，在激素/神经递质的产生中发挥重要作用；描述了一个详细的程序，该程序检查了与C-H和氧激活过程有关的蛋白质的关键区域。铜酶半乳糖氧化酶(Galo)被认为产生类似的铜(II)-超氧中间体，将作为理解O-H与C-H激活的基准与T2M进行对比。2-组氨酸，1-天冬氨酸家族的非血红素酶利用一个共同的活性部位来功能化一系列不同的反应。对这一家族中依赖不同还原剂的两个成员进行了研究：1-氨基环丙烷1-羧酸氧化酶(ACCO)反应中的抗坏血酸和牛磺酸双加氧酶(TauD)反应中的α-酮戊二酸。TauD的目标是详细研究隧道效应，而ACCO的研究将集中在ACC击穿的谜团路径及其与氧激活的联系上。最后，铜胺氧化酶(CAO)催化氢离子和葡萄糖氧化酶(GO)催化氢离子脱除带电氢离子的两种酶系统为研究底物专一性(CAO)和反应驱动力(GO)之间的相互关系提供了独特的机会。上述研究对我们理解自然的催化剂具有基本意义，以前发表的本实验室的发现导致了关于酶催化的物理起源的范式转变。对后者的理解为合成小型仿生催化剂、设计新型蛋白质/多肽催化剂以及建立指导药物设计的概念性平台提供了指导。与公共健康相关有氧生命的两个最基本的过程是C-H和O2的激活。本申请的目的是阐明和编纂支配这些反应的酶激活的原理。

DESCRIPTION (provided by applicant): The overarching aim of this application is to elucidate the principles that govern enzyme catalyzed C-H and O2 activation processes. Studies of C-H activation (proton, hydride and hydrogen atom) are focused on understanding the role of quantum mechanical hydrogen transfer and its link to protein structure and dynamics. The investigation of oxygen activation is directed toward reactions that remain particularly enigmatic with regard to the nature of the oxygen species and/or the reaction pathway that leads to product formation. The pharmacologically important enzyme lipoxygenase has become a center pin of studies aimed at understanding the role of hydrogen tunneling in enzyme catalysis; future studies include probes of protein motions and their links to the tunneling event, the impact of high pressure on tunneling, and the characterization of a thermophilic homolog for comparison to the mesophilic soybean lipoxygenase. The channeling of oxygen within a discrete region of protein, orthogonal to the substrate-binding pocket, has been postulated in lipoxygenase; this will be further tested using site-specific mutagenesis. Tyramine ?- monooxygenase (T2M) is a member of a small family of eukaryotic copper proteins that play essential roles in the generation of hormones/neurotransmitters; a detailed program is described that examines key regions of the protein in relation to the C-H and oxygen activation processes. The copper enzyme galactose oxidase (GalO), postulated to generate a similar Cu(II)-superoxo- intermediate, will be contrasted to T2M as a benchmark for understanding O-H vs. C-H activation. The 2-His, 1-Asp family of non-heme enzymes utilizes a common active site to functionalize a wide range of disparate reactions. Studies are described to examine two members of this family that depend on different reductants; ascorbate in the 1-aminocyclopropane 1- carboxylate oxidase (ACCO) reaction and alpha-ketoglutarate in the taurine dioxygenase (TauD) reaction. TauD is targeted for detailed studies of tunneling, while studies of ACCO will focus on the enigmatic pathway for ACC breakdown and its link to oxygen activation. Finally, two enzyme systems that catalyze removal of a charged hydrogen species as a proton, catalyzed by copper amine oxidase (CAO), and as a hydride ion, catalyzed by glucose oxidase (GO), provide unique opportunities to interrogate the inter- relationship between substrate specificity (CAO) and reaction driving force (GO) in the tunneling process. The above studies are of fundamental significance to our understanding of Nature's catalysts, with previously published findings from this laboratory leading to paradigm shifts regarding the physical origins of enzyme catalysis. Understanding the latter provides guidelines for the synthesis of small biomimetic catalysts, for the design of novel protein/peptide based catalysts and for the establishment of conceptual platforms that guide drug design. PUBLIC HEALTH RELEVANCE Two of the most fundamental processes that underlie aerobic life are C-H and O2 activation. The goal of this application is to elucidate and codify the principles that govern enzyme activation of these reactions.