Three-membered Ring Metabolites, Inhibition and Formation

描述(申请人提供)：许多天然产品含有作为关键结构元素的环丙烷、环氧乙烷或氮杂环丁基团。这些三元环部分通常是稳定的，尽管它们有相当大的环张力。然而，这些小环的固有反应性可以通过酶激活来释放。这种激活通常会导致活性中间体抑制相应的酶，使这些含有三元环的化合物成为潜在的药物。尽管这些小的环状结构作为治疗剂和机械探针有很长的历史，但人们对它们在自然界中是如何构建的知之甚少。为了探索这些应变环化合物的生物合成并促进药物设计工作，我们选择了研究几种与环氧乙烷和氮杂环丙烷形成有关的有趣的酶。其中包括磷霉素生物合成途径中的(S)-2-羟丙基膦酸环氧酶(HppE)和2-羟乙基膦酸甲基转移酶(HEPM)，以及阿奇霉素A生物合成途径中催化氮杂环形成的酶。这些酶之所以被选中，是因为它们具有重要的生物学作用，它们新颖的催化机制，以及它们作为新疗法组合生物合成催化剂的潜力。拟议的实验将针对以下特定目标：(1)研究HppE的催化机理，包括反应中间产物的氧活化机理和化学性质；(2)表征HEM的催化性能，特别是甲钴胺和自由基-SAM在催化中的功能，以及甲基化反应的机理；(3)建立阿奇霉素A中氮杂环丙烷的生物合成途径，并表征参与转化的关键酶。这些研究不仅有助于更好地了解靶标酶的催化机理，还将为设计控制和模拟相关酶的催化功能的方法提供重要的见解，其中许多酶具有医学意义。我们的研究结果有望为天然产物生物合成和机械酶学的广泛领域做出贡献，也可能为未来利用途径工程和/或组合生物合成方法开发新代谢物的临床应用提供帮助。与公众健康相关：这项应用的目标是了解含氧和含氮的三元环化合物是如何生物合成的，以及涉及的关键酶的机制。这项工作所获得的见解将有助于开发具有治疗潜力的新的小环制剂，从而对人类健康产生积极的影响。

DESCRIPTION (provided by applicant): Many natural products contain cyclopropane, oxirane, or aziridine groups as key structural elements. These three-membered ring moieties are generally stable despite their considerable ring strain. However, the inherent reactivities of these small rings can be released by enzymatic activation. Such activation often leads to reactive intermediates that inhibit the corresponding enzyme, making these three-membered ring containing compounds potential drugs. Although these small ring structures have a long history as therapeutic agents and mechanistic probes, little is known about how they are constructed in nature. To explore the biosynthesis of these strained ring compounds and to facilitate drug design efforts, we have chosen to study several intriguing enzymes involved in oxirane and aziridine formation. These include (S)-2- hydroxypropylphosphonate epoxidase (HppE) and 2-hydroxyethylphosphonate methyltransferase (HepM) in the fosfomycin biosynthetic pathway, and the enzymes catalyzing aziridine ring formation in the azicemicin A biosynthetic pathway. These enzymes were selected for their significant biological roles, their novel catalytic mechanisms, and their potential as catalysts for the combinatorial biosynthesis of new therapeutics. The proposed experiments will address the following specific aims: (1) to investigate the catalytic mechanism of HppE, including the oxygen activation mechanism and the chemical nature of the reaction intermediates; (2) to characterize the catalytic properties of HepM, especially the functions of methylcobalamin and radical- SAM in catalysis, and the mechanism of the methylation reaction; (3) to establish the biosynthetic pathway of aziridine ring formation in azicemicin A, and to characterize the key enzymes involved in the transformation. These studies will not only lead to a better understanding of the catalytic mechanism of the targeted enzymes, but will also provide important insight for designing methods to control and mimic the catalytic functions of related enzymes, many of which are medically relevant. Our results are expected to contribute to the broad field of natural product biosynthesis and mechanistic enzymology, and may also assist future clinical applications for the development of new metabolites using pathway engineering and/or combinatorial biosynthetic methods. PUBLIC HEALTH RELEVANCE: The objective of this application focuses on learning how oxygen and nitrogen containing three-membered ring compounds are biosynthesized and the mechanisms of the key enzymes involved. The insight gained from this work will be useful for the development of new small ring agents having therapeutic potential, and thus will have a positive impact on human health.