Accessing and Expanding Natural Products Chemical Diversity by Big-data Analysis and Biosynthetic Investigation

项目摘要/摘要 天然产物(NPs)历史上一直是生物活性分子的重要来源，NPs和它们的 衍生品占FDA批准的小分子药物的50%以上。近年来，以NP为基础的药物 由于重复发现相同或相同的药物，发现面临着识别新药的根本障碍 类似的化合物，代表有限的化学多样性。幸运的是，由于NPs一直在进化 数十亿年来，在千万亿不同的环境中，有大量新的生物活性NPs被编码 在自然界中，这可能是有用的药物。然而，它们的可及性是一个问题：只有不到10%的NP 生物合成基因簇(BGC)已经与现有的NPs相连，留下了绝大多数BGC 关于它们可能产生什么NPs的问题还没有得到开发。该研究计划的总体目标是利用大数据 信息分析和生物合成研究，以获取和转化巨大的遗传潜力 这些“孤儿BGC”，即具有未知产品的BGC，进入化学现实，将它们与他们的产品和 进而为药物发现筛选提供结构多样化的NPs池。为此，我们提出了两项建议 研究方向：(1)利用我们建立的大数据关联网络分析，我们已经确定 几乎所有III类羊毛肽的BGC中都缺少隐藏的蛋白酶。我们以前使用过这种方法 首次从菲尔米库斯中发现两个新的III类羊硫肽家族。我们将利用这些优势 隐藏的蛋白酶，以进一步解开羊毛肽固有的化学多样性，并生成两个 天然和非天然多肽的体外酶促合成和靶向生物合成工程 药物发现筛选。(2)挖掘尚未开发的微生物遗传潜力，初步强调硫- 包含NPs和前所未有的生物合成途径杂交，我们优先考虑了两个有前途的孤儿 BGC具有高度独特的酶学特性，并将它们与其本地产品连接起来。第一部以小说S为特色-- 羟化黄素蛋白，可能参与形成新的含硫官能团。这个 第二个具有史无前例的萜类-脂肪酸-非核糖体多肽杂交由不同寻常的 交叉途径的酶组合。我们将进一步研究它们所蕴含的新的生物合成 BGC将产生新的NPs，为未来类似途径的基因组挖掘提供信息，并使途径工程成为可能 以进一步增加NPs的化学多样性。我们在这两个研究方向上的重大进展支持了 这项建议的可行性以及我们建立一个成功和可持续的独立 在这一领域的计划。我们在生物活性筛选和蛋白质结构方面促进了几个关键的合作 生物学进一步加强了我们的研究计划。此外，该项目还将提供培训机会 本科生、毕业生和博士后研究员。总体而言，该计划有望发现新的 生物合成，扩大NPs化学多样性，促进基于信息学的NPs发现和生物工程 以提供有希望的新药线索。

PROJECT SUMMARY/ABSTRACT Natural products (NPs) have historically been a critical source of bioactive molecules, with NPs and their derivatives making up over 50% of FDA-approved small molecule drugs. In recent years, NP-based drug discovery is facing a fundamental barrier in identifying new drugs due to repeated rediscovery of the same or similar compounds, representing limited chemical diversity. Fortunately, since NPs have been evolving over billions of years in trillions of vastly diverse environments, there is an abundance of new bioactive NPs encoded in nature which may be useful as drugs. However, their accessibility is a problem: only less than 10% of NP biosynthetic gene clusters (BGCs) have been connected to existing NPs, leaving the vast majority of BGCs untapped as to what NPs they may produce. The overall goal of this research program is to leverage big-data informatic analysis and biosynthetic investigation to access and convert the tremendous genetic potential of these “orphan BGCs”, BGCs with unknown products, into chemical reality, connecting them to their products and in turn supplying structurally diverse pools of NPs for drug discovery screening. To this end, we propose two research directions: (1) Utilizing our established big-data correlational networking analysis, we have identified hidden proteases missing from the BGCs of almost all class III lanthipeptides. We previously used this method to discover two new families of class III lanthipeptides from Firmicutes for the first time. We will leverage these hidden proteases to further unlock the inherent chemical diversity of lanthipeptides and generate two libraries of natural and non-natural peptides through in vitro enzymatic synthesis and targeted biosynthetic engineering for drug discovery screening. (2) Mining the untapped microbial genetic potential, with an initial emphasis on sulfur- containing NPs and unprecedented biosynthetic pathway hybridization, we have prioritized two promising orphan BGCs with highly unique enzymology and connected them to their native products. The first features a novel S- hydroxylating flavoprotein, potentially involved in the formation of a new sulfur-containing functionality. The second has an unprecedented terpenoid-fatty acid-non-ribosomal peptide hybridization mediated by unusual cross-pathway enzymatic combinations. We will further investigate the new biosynthesis harbored by these BGCs to produce new NPs, inform future genome mining of similar pathways, and enable pathway engineering to further increase NPs chemical diversity. Our significant progress in both research directions supports the feasibility of this proposal as well as our competence to establish a successful and sustainable independent program in this field. We have fostered several key collaborations in bioactivity screening and protein structural biology that further strengthen our research program. In addition, this program will provide opportunities to train undergraduates, graduates, and postdoctoral fellows. Overall, this program is expected to discover new biosynthesis, expand NPs chemical diversity, and facilitate informatics-based NPs discovery and bioengineering to provide promising new drug leads.