Interrogation of Microbial Natural Product Methyltransferases

微生物天然产物甲基转移酶的研究

• 批准号: 10674546
• 负责人: Tony D. Davis
• 金额: $ 24.9万
• 依托单位: STATE UNIVERSITY OF NY,BINGHAMTON
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-15 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10674546
• 关键词: Affinity; Anabolism; Anti-Bacterial Agents; Antibiotics; Binding; Biological; Carrier Proteins; Catalysis; Cell physiology; Cells; Clinic; Communication; Detection; Drug Design; Drug Targeting; Engineering; Enzymes; Epigenetic Process; Eukaryota; Fostering; Future; Gene Expression Regulation; Goals; In Vitro; Individual; Infection; Invaded; Investigation; Life; Mediating; Methyltransferase; Molecular; Molecular Biology; Multiple Bacterial Drug Resistance; Mutagenesis; Natural Products; Nature; Neurons; Nucleic Acids; Organic Synthesis; Pathogenicity; Pathway interactions; Peptides; Pharmaceutical Chemistry; Pharmacologic Substance; Play; Prevalence; Production; Productivity; Property; Protein Engineering; Protein Methyltransferases; Proteins; Proteome; Research; Resistance; Role; S-Adenosylhomocysteine; S-Adenosylmethionine; Signal Transduction; Signaling Molecule; Specificity; Structure; Substrate Specificity; Virulence Factors; Work; activity-based protein profiling; alkyl group; analog; anti-cancer; antimicrobial drug; biophysical techniques; cofactor; combinatorial; cost effective; design; in vivo; inhibitor; insight; microbial; microorganism; natural product derivative; novel; nucleic acid methylation; pathogen; pharmacophore; polyketides; pressure; programs; protein complex; protein protein interaction; rational design; screening; small molecule; structural biology; therapeutic target; tool

PROJECT SUMMARY/ABSTRACT The increased prevalence of multi-drug resistant bacteria has led to an urgent unmet need to develop new an- timicrobial drugs. Targeting bacterial natural product biosynthesis has emerged as a promising avenue, due to the role of natural products as virulence factors, signaling factors, and agents of microbial warfare. However, to date, no drugs targeting enzymes in these biosynthetic pathways have reached the clinic. Hence, systematic investigations into key biosynthetic enzymes are necessary to understand fully their individual roles in virulence factor production. S-adenosyl-L-methionine (SAM)-dependent methyltransferases are ubiquitous throughout all domains of life and modify a diverse array of substrates, including proteins, nucleic acids, and natural products. In eukaryotes, methyltransferase activity modulates cellular processes such as epigenetic gene regulation and neuronal communication; consequently, these methyltransferases are extensively studied and have been validated as anticancer targets. In contrast, natural product methyltransferases have yet to be explored as therapeutic “tar- gets”, in spite of their important role in the biosynthesis of pathogenic virulence factors and pharmaceutically relevant compounds. During biosynthesis, natural product methyltransferases recognize substrates that are covalently tethered to a carrier protein; hence, successful methyl transfer is a function of both substrate recog- nition and proper protein-protein interactions between the methyltransferase and the carrier protein. The tran- sient nature of these protein-protein interactions makes targeting methyltransferases particularly challenging, yet elucidating the molecular basis of cofactor-protein, substrate-protein, and protein-protein recognition would offer a significant step towards advancing these efforts. Herein, our overall goal is to identify and engineer critical methyltransferase interactions during the biosynthe- sis of non-ribosomal peptide and polyketide-derived virulence factors by combining organic synthesis, ad- vanced molecular biology, and structural biology. We propose to (aim 1) develop small molecules to interro- gate cofactor and substrate methyltransferase activity, (aim 2) design tools to capture methyltransferase-carrier protein interactions, and (aim 3) develop selective activity-based probes for bacterial natural product methyl- transferases for detection and identification in the bacterial proteome. The insights from this work will be broad- ly significant by providing critical first steps in structure-based design of drugs targeting bacterial methyltrans- ferases, fostering combinatorial biosynthetic efforts of alkyl groups into unnatural products with precise regio-, stereo-, and chemoselectivity, and advancing medicinal chemistry efforts that alter biological and physico- chemical properties of privileged pharmacophores through chemoenzymatic synthesis.

项目总结/摘要 多重耐药细菌的流行增加导致了开发新的抗- 抗菌药物靶向细菌天然产物生物合成已成为一种有前途的途径，这是由于 天然产物作为毒力因子、信号因子和微生物战剂的作用。但要 迄今为止，还没有靶向这些生物合成途径中的酶的药物到达临床。因此，系统 对关键的生物合成酶的研究对于充分了解它们在毒力中的作用是必要的 生产要素。 S-腺苷-L-甲硫氨酸（SAM）依赖性甲基转移酶在生命的各个领域都是普遍存在的 并修饰多种底物，包括蛋白质、核酸和天然产物。在真核生物中， 甲基转移酶活性调节细胞过程，如表观遗传基因调控和神经元 因此，这些甲基转移酶被广泛研究，并已被验证为 抗癌靶点相比之下，天然产物甲基转移酶尚未被探索作为治疗性“焦油”。 尽管它们在致病性毒力因子的生物合成和药物治疗中起重要作用， 相关化合物。在生物合成过程中，天然产物甲基转移酶识别底物， 共价连接到载体蛋白;因此，成功的甲基转移是两个底物识别的功能， 甲基转移酶和载体蛋白之间的启动和适当的蛋白质-蛋白质相互作用。trans- 这些蛋白质-蛋白质相互作用的沉默性质使得靶向甲基转移酶特别具有挑战性， 然而，阐明辅因子-蛋白、底物-蛋白和蛋白-蛋白识别的分子基础将 为推进这些努力迈出了重要一步。 在此，我们的总体目标是确定和工程关键甲基转移酶的相互作用，在生物合成过程中， 非核糖体肽和聚酮衍生毒力因子通过有机合成、ad- 先进的分子生物学和结构生物学。我们建议（目标1）开发小分子， 门辅因子和底物甲基转移酶活性，（目的2）设计捕获甲基转移酶载体的工具 蛋白质相互作用，并（目的3）开发基于活性的细菌天然产物甲基- 用于细菌蛋白质组中检测和鉴定的转移酶。从这项工作的见解将是广泛的- 通过提供关键的第一步，在基于结构的药物设计靶向细菌甲基反式， ferases，促进烷基的组合生物合成的努力，以精确的区域， 立体，和化学选择性，并推进药物化学的努力，改变生物和物理， 通过化学酶促合成的特权药效团的化学性质。