Mechanisms of Biosynthetic Formation of Deoxy Sugars

说明(申请人提供)：许多具有生物活性的天然产物的活性来源于其结构中的糖成分。改变这些糖的结构可以对母体化合物的生物活性、选择性和药代动力学特性产生深远的影响。这一观察结果推动了利用糖生物合成机制来衍生各种含糖量不同的天然产品的方法的发展。充分认识这种方法的潜力依赖于新的糖生物合成途径的发现，还需要彻底了解每一种目标糖的生物合成途径，包括关键酶的详细机制知识。考虑到这些目标，我们在以前的赠款资助的工作中取得了显著的成果。根据这些研究的结果，我们确定了四个需要在下一个资金阶段进一步研究的主要领域。因此，本申请概述了旨在了解脱糖胺、apiose、kijanose和2-脱氧-2-巯基葡萄糖是如何生物合成的实验。具体目标包括(1)参与许多大环内酯类抗生素的重要成分去糖胺生物合成的自由基-SAM酶(DesII)的机理研究；(2)UDP-apiose合成酶催化吡喃糖到呋喃糖环收缩的机制的确定；(3)对KijAnimicin中不寻常的硝糖部分(Kijanose)的生物合成的研究；以及(4)抗生素BE-7585A的2-脱氧-2-巯基葡萄糖部分中硫结合的机制的阐明。拟议的实验不仅将描绘脱氧、支链、硝基和含硫糖的生物合成，而且还将通过增进我们对几类重要酶的理解来推进机械酶学领域。我们的结果对应用生物医学研究也是有价值的，因为新的糖基化工具(即糖的生物合成途径和酶)将被发现用于未来的次生代谢物糖多样化努力。与公众健康相关：在本申请中概述了旨在研究自由基SAM酶DesII参与地塞米胺生物合成中C4-脱氧步骤的机制、UDP-apiose合成酶(Axs1)催化的环收缩反应的机制、一种不寻常的硝糖-Kijanose的生物合成以及BE-7585A的硫糖部分掺入硫的机制的实验。这一结果有望显著推进机械酶学领域的发展，并对应用生物医学研究具有重要价值，因为将为未来的次生代谢物糖多样化努力发现新的糖生物合成途径和酶。

DESCRIPTION (provided by applicant): Many biologically active natural products derive their activity from the sugar components of their structures. Changing the structures of these sugars can have a profound impact on the biological activity, selectivity, and pharmacokinetic properties of the parent compounds. This observation has fueled the development of methods to derivatize natural products with diverse sugar moieties by exploiting the sugar biosynthetic machinery. Fully realizing the potential of such an approach relies on the discovery of new sugar biosynthetic pathways, and also requires a thorough understanding of the biosynthetic pathway of each target sugar including detailed mechanistic knowledge of the key enzymes. With these goals in mind, we have produced notable results in work funded by previous grant. As a result of these studies, we have identified four key areas that warrant further investigation in the next funding period. Accordingly, this application outlines experiments designed to learn how desosamine, apiose, kijanose, and 2-deoxy-2- mercaptoglucose are biosynthesized. The specific objectives include (1) mechanistic studies of a radical-SAM enzyme (DesII) involved in the biosynthesis of desosamine, an essential component of many macrolide antibiotics, (2) a determination of the mechanism of the pyranose-to-furanose ring-contraction catalyzed by UDP-apiose synthase, (3) an investigation into the biosynthesis of the unusual nitrosugar moiety (kijanose) of kijanimicin, and (4) the elucidation of the mechanism of sulfur incorporation into the 2-deoxy-2-mercaptoglucose moiety of the antibiotic BE-7585A. The proposed experiments will not only delineate the biosynthesis of deoxy-, branched-chain, nitro-, and sulfur-containing sugars, but will also advance the field of mechanistic enzymology by enhancing our understanding of several important classes of enzymes. Our results should also be valuable to applied biomedical research, as new glycosylation tools (i.e., sugar biosynthetic pathways and enzymes) will be discovered for future secondary metabolite glycodiversification efforts. PUBLIC HEALTH RELEVANCE: Outlined in this application are experiments designed to study the mechanism of the radical SAM enzyme, DesII, involved in the C4-deoxygenation step in desosamine biosynthesis, the mechanism of the ring contraction reaction catalyzed by UDP-apiose synthase (Axs1), the biosynthesis of an unusual nitrosugar, kijanose, and the mechanism of sulfur incorporation into the thiosugar moiety of BE-7585A. The results are expected to significantly advance the field of mechanistic enzymology and should also be valuable to applied biomedical research, as new sugar biosynthetic pathways and enzymes will be discovered for future secondary metabolite glycodiversification efforts.