Chemical and biological exploration of a new natural product family, the thiazole

新天然产物家族噻唑的化学和生物学探索

• 批准号: 8077660
• 负责人: Douglas Alan Mitchell
• 金额: $ 25.76万
• 依托单位: UNIVERSITY OF ILLINOIS AT URBANA-CHAMPAIGN
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-02-01 至 2016-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8077660
• 关键词: Address; Anabolism; Anti-Infective Agents; Antibiotics; Area; Bacillus (bacterium); Bacillus cereus; Bacteria; Binding; Biological; Biological Assay; Biological Factors; Biological Process; Calorimetry; Catalysis; Cellular Membrane; Chemical Structure; Chemicals; Complex; Comprehension; DNA Gyrase; Data; Databases; Development; Docking; Enzymes; Exhibits; Family; Figs - dietary; Foundations; Fourier Transform; Frequencies; Funding; Future; Gene Deletion; Genes; Genetic; Genome; Genomics; Human; In Vitro; Kinetics; Knowledge; Laboratories; Lead; Ligase; Mass Spectrum Analysis; Medicine; Metabolism; Methods; Microbe; Mining; Modern Medicine; Molecular; Names; Nature; Organism; Oxazoles; Oxidoreductase; Pathogenesis; Pathway interactions; Peptides; Pharmaceutical Preparations; Pharmacologic Substance; Plants; Play; Prize; Property; Proteins; Research; Resolution; Ribosomes; Roentgen Rays; Role; Secondary to; Site-Directed Mutagenesis; Soil; Solid; Source; Specificity; Spectrum Analysis; Structure; Surface; Surface Plasmon Resonance; Therapeutic Agents; Thiazoles; Thiostrepton; TimeLine; Toxin; Ursidae Family; Virulence; Vision; Work; arm; base; combinatorial; inhibitor/antagonist; insight; interest; member; microbial; microcin; novel; novel therapeutics; pathogen; programs; protein protein interaction; reconstitution; small molecule; streptolysin S; success; tool; weapons

DESCRIPTION (provided by applicant): The genomics revolution has repeatedly demonstrated that our understanding of natural product (NP) biosynthesis is far from complete. Given the frequency of silent and cryptic biosynthetic clusters in existing and emerging genomes, compounded with the inability to culture >99% of microbes, far less than 1% of microbial NPs have been discovered. This project proposes to characterize novel NP biosynthetic clusters from two soil-dwelling bacteria. Without question, NPs from soil bacteria are our most prolific source of medicine. Unlocking the chemical structure and biological function of novel NPs encoded by these organisms holds enormous potential for expanding our pharmaceutical repertoire. The biosynthetic clusters of interest to this proposal are members of a recently described, evolutionarily conserved family dubbed the thiazole/oxazole-modified microcins (TOMM). As a new NP family, TOMMs represent an underexplored area of NP chemical space - few have a known structure or mechanism of action. All TOMMs with a known activity function as toxins, making them of paramount interest to modern medicine. In two known cases, TOMMs produced by human pathogens play a critical role in the molecular mechanism of pathogenesis. Therefore, a more complete knowledge of the biosynthetic pathway could lead to the development of virulence-targeting antibiotics, which represents a longer-term objective for our research program. To effectively tap into this potential, several gaps in our current understanding of these molecules must be addressed. This project is divided into three related, but independent specific aims. For Aim 1, a combination of in vitro reconstitution, natural product isolation, and advanced spectroscopy will be employed to determine the chemical structure of the TOMM product. In Aim 2, high-resolution mass spectrometry and site-directed mutagenesis will be used to kinetically evaluate a key enzyme that catalyzes the first step in the formation of thiazoles and oxazoles. This enzyme, a cyclodehydratase, is responsible for recognizing the TOMM precursor peptide and converting Cys and Ser/Thr residues into thiazolines and (methyl) oxazolines. Aim 3 seeks to reveal the protein-protein interactions that enable substrate recognition and the downstream thiazole/oxazole forming activity. By characterizing the enzymes involved in TOMM biosynthesis, the foundation for future work will be laid, including the development of biosynthetic inhibitors of TOMMs from human pathogens and strategies to harness the power of combinatorial biosynthesis to evolve TOMMs with desired biological targets. Progress on this project will fill a major void in our current understanding of how a subset of peptide-derived toxins is biosynthesized. The tools developed will be broadly applicable to the study of other TOMMs. PUBLIC HEALTH RELEVANCE: The majority of medicines used today are derived from complex chemicals produced by nature. Such natural products continue to be our most valuable source of new drugs. To lay the foundation for expanding the current repertoire of pharmaceuticals, we seek to characterize how bacteria synthesize a new subclass of natural products.

描述（由申请人提供）：基因组革命反复证明我们对天然产物（NP）生物合成的理解远非完整。鉴于现有基因组和新兴基因组中静音和隐性生物合成簇的频率，与无法培养> 99％的微生物相比，已经发现了少于1％的微生物NP。该项目提出了表征来自两个土壤细菌的新型NP生物合成簇。毫无疑问，来自土壤细菌的NP是我们最多产的药物来源。解锁由这些生物编码的新型NP的化学结构和生物学功能具有扩展我们的药物库的巨大潜力。该提案感兴趣的生物合成簇是最近描述的，进化保守的家族的成员，称为噻唑/恶唑修饰的微蛋白（TOMM）。作为一个新的NP家族，Tomms代表了NP化学空间的一个未充分倍增的区域 - 很少有已知的结构或作用机理。所有具有已知活动功能的毒素的Tomms，使其对现代医学具有最高兴趣。在两种已知情况下，人类病原体产生的汤点在发病机理的分子机理中起关键作用。因此，对生物合成途径的更完整的了解可能会导致靶向毒力的抗生素的发展，这代表了我们的研究计划的长期目标。为了有效利用这一潜力，我们目前对这些分子的理解中的几个差距必须解决。该项目分为三个相关但独立的特定目的。对于AIM 1，将使用体外重构，天然产物隔离和晚期光谱法的结合来确定TOMM产物的化学结构。在AIM 2中，高分辨率的质谱法和位置指导的诱变将用于评估一种关键酶，该酶催化了噻唑和氧唑形成的第一步。该酶是一种环氧化氢蛋白酶，负责识别TOMM前体肽，并将Cys和Ser/Thr残基转化为噻唑啉和（甲基）恶作剂。 AIM 3试图揭示蛋白质 - 蛋白质相互作用，从而使底物识别和下游噻唑/恶唑形成活性。通过表征参与Tomm Biosynsess的酶，将奠定未来工作的基础，包括从人类病原体和策略中开发Tomms的生物合成抑制剂，以利用联合生物合成的力量，从而以所需的生物学靶标的进化Tomms。该项目的进展将填补我们当前对肽衍生毒素的子集如何生物合成的主要空隙。开发的工具将广泛适用于其他Tomms的研究。 公共卫生相关性：当今使用的大多数药物均来自自然产生的复杂化学物质。这种天然产品仍然是我们最有价值的新药来源。为了奠定扩大当前药品库的基础，我们试图表征细菌如何合成新的天然产品子类。