Three-membered Ring Metabolites, Inhibition and Formation

描述（由申请人提供）：许多天然产物含有环丙烷、氧环烷或叠氮吡啶基团作为关键结构元素。尽管这些三元环具有相当大的环应变，但它们通常是稳定的。然而，这些小环的固有反应性可以通过酶激活释放出来。这种活化通常会导致活性中间体抑制相应的酶，使这些含三元环的化合物成为潜在的药物。尽管这些小环结构作为治疗剂和机械探针已有很长的历史，但人们对它们在自然界中的构造知之甚少。为了探索这些张力环化合物的生物合成并促进药物设计工作，我们选择了研究一些有趣的酶，这些酶参与了氧环和氮吡啶的形成。这些酶包括磷霉素生物合成途径中的(S)-2-羟丙基膦酸酯环氧化酶（HppE）和2-羟乙基膦酸酯甲基转移酶（HepM），以及azicemicin A生物合成途径中的催化叠氮环形成的酶。这些酶之所以被选中，是因为它们具有重要的生物学作用、新的催化机制以及作为新疗法组合生物合成催化剂的潜力。本实验将针对以下具体目标：(1)研究HppE的催化机理，包括氧活化机理和反应中间体的化学性质；(2)表征HepM的催化性能，特别是甲基钴胺素和自由基- SAM在催化中的作用，以及甲基化反应的机理；(3)建立叠氮霉素A形成叠氮环的生物合成途径，并对参与转化的关键酶进行表征。这些研究不仅将有助于更好地了解目标酶的催化机制，而且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见解，其中许多与医学相关。我们的研究结果有望为天然产物生物合成和机制酶学的广泛领域做出贡献，并可能有助于未来利用途径工程和/或组合生物合成方法开发新的代谢物的临床应用。公共卫生相关性：此应用程序的目标侧重于了解含氧和氮的三元环化合物是如何生物合成的以及所涉及的关键酶的机制。从这项工作中获得的见解将有助于开发具有治疗潜力的新型小环药物，从而对人类健康产生积极影响。

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.