Anthranilate-Based Fungal Nonribosomal Peptide Synthetase Assembly Lines

基于邻氨基苯甲酸的真菌非核糖体肽合成酶装配线

• 批准号: 7803244
• 负责人: Brian Douglas Ames
• 金额: $ 4.56万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-01-01 至 2012-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7803244
• 关键词: Acyltransferase; Affinity; Alkaloids; Amino Acids; Anabolism; Ants; Aspergillus; Aspergillus fumigatus; Bacteria; Benzodiazepines; Binding; Biochemical; Biochemistry; Bioinformatics; Biological; Biological Assay; Biological Factors; Biology; Brain; C-terminal; Characteristics; Chemicals; Chemistry; Cholecystokinin Receptor; Cloning; Coenzyme A; Collaborations; Crystallography; Cyclization; Data; Drug Design; Engineering; Environment; Enzymatic Biochemistry; Enzymes; Event; Fermentation; Figs - dietary; Foundations; Fungal Genome; Fungal Proteins; Gastrointestinal tract structure; Gene Cluster; General Hospitals; Generations; Genes; Genome; Homology Modeling; In Vitro; Investigation; Knowledge; Literature; Mass Spectrum Analysis; Massachusetts; Mediating; Methods; Mining; Molecular; Molecular Cloning; NMR Spectroscopy; Nature; Neuraxis; Oxygenases; Pathway interactions; Pattern; Peptides; Pharmacology; Physical condensation; Polycyclic Compounds; Process; Protein Isoforms; Publications; Publishing; Quinazolines; Reporting; Research; Research Personnel; Resources; Role; Route; Sequence Analysis; Solid; Structure; System; Targeted Toxins; Therapeutic; Therapeutic Agents; Time; Toxin; Universities; Validation; Vertebral column; anthranilate; asperlicin; base; chemical synthesis; enzyme activity; experience; feeding; fumiquinazoline A; fungus; gastrointestinal; insight; medical schools; nervous system disorder; peptide synthase; plant fungi; professor; protein expression; protein structure; public health relevance; reconstitution; research study; scaffold; small molecule; tool

DESCRIPTION (provided by applicant): The application's broad objective is to provide new molecular insights into the biosynthetic assembly of medicinally relevant fungal polycyclic compounds by nonribosomal peptide synthetases (NRPSs). Two major classes of amino acid-derived multicyclic fungal metabolites are the benzodiazepinones and quinazolinones. Core structures are derived from combination of the beta amino acid anthranilate (ortho-aminobenzoate) with proteinogenic amino acids, resulting in the formation of scaffolds with potent bioactivities, both as therapeutics (such as the asperlicins) and toxins (such as acetylaszonalenin). The anthranilate-containing fungal metabolite asperlicin is a cholecystokinin (CCK) receptor antagonist of nanomolar potency. CCK receptor isoforms are found in the brain, central nervous system and alimentary canal; CCK receptor antagonists provide a route to treat both gastrointestinal and neurological disorders. The NRPS-based morphing of linear peptide chains into architecturally constrained fused-ring systems with diverse substituents underlies the high affinities and biological selectivities observed for these classes of compounds. Previous studies have provided structures and synthetic routes for many compounds of these classes, but little is known regarding their biosynthesis beyond building block composition through precursor feeding studies. The proposed research aims to provide a fundamental understanding of: 1) the selection, activation, and loading of anthranilate by fungal NRPSs, and 2) the chemical processes mediated by the biosynthetic enzymes to cyclize and tailor the linear peptide into the final bioactive product. We predict that the number and order or anthranilate building blocks incorporated into the peptide backbone are key determinants to multicylic scaffold construction. Completion of the proposed research will provide knowledge that may be applied to the directed incorporation of anthranilate into peptide backbones to engineer non-natural cyclization patterns and substituents. Through genome mining, biochemical experimentation, and structural analysis we expect to characterize the molecular basis of anthranilate selection and loading by NRPSs. Through deconstruction and reconstitution of the biosynthetic machinery activity, we will explore a predicted new mode of NRPS-based cyclization of the peptide backbone for multicylic scaffold construction. PUBLIC HEALTH RELEVANCE: Understanding the biological assembly of anthranilate-containing fungal natural products will provide a foundation for producing molecules to screen as therapeutic agents, and will identify biological targets from toxin producing pathways for drug design.

描述（由申请人提供）：申请的主要目标是通过非核糖体肽合成酶（NRPSs）为药用真菌多环化合物的生物合成组装提供新的分子见解。两大类氨基酸衍生的多环真菌代谢产物是苯二氮卓酮类和喹唑啉酮类。核心结构来源于-氨基酸邻氨基苯甲酸酯（邻氨基苯甲酸酯）与蛋白质原氨基酸的结合，从而形成具有强大生物活性的支架，既可以作为治疗药物（如曲霉毒素），也可以作为毒素（如乙酰丙烯酰胺）。含有邻氨基苯甲酸的真菌代谢物曲霉素是一种纳米摩尔效价的胆囊收缩素受体拮抗剂。CCK受体亚型存在于大脑、中枢神经系统和消化道；CCK受体拮抗剂提供了一种治疗胃肠道和神经系统疾病的途径。基于nrps的线性肽链变形成具有不同取代基的结构约束的融合环系统，这是观察到的这类化合物的高亲和力和生物选择性的基础。以往的研究已经提供了许多这类化合物的结构和合成途径，但对它们的生物合成知之甚少，除了通过前体取食研究构建块组成。本研究旨在为以下方面提供基础知识：1)真菌NRPSs对邻氨基苯甲酸酯的选择、激活和负载；2)生物合成酶介导线性肽环化和裁剪成最终生物活性产物的化学过程。我们预测纳入肽骨架的邻氨基甲酸酯构建块的数量和顺序是多环支架构建的关键决定因素。该研究的完成将提供可应用于将邻氨基苯甲酸酯直接掺入肽骨架以设计非自然环化模式和取代基的知识。通过基因组挖掘，生化实验和结构分析，我们期望表征NRPSs对邻氨基苯甲酸酯选择和装载的分子基础。通过对生物合成机械活性的解构和重构，我们将探索一种基于nrps的多肽主链环化的预测新模式，用于构建多环支架。