Development of a Bacterial Host for Natural Product Discovery and Production

用于天然产物发现和生产的细菌宿主的开发

• 批准号: 10263933
• 负责人: Alessandra S Eustaquio
• 金额: $ 36.96万
• 依托单位: UNIVERSITY OF ILLINOIS AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-20 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10263933
• 关键词: Address; Anabolism; Autologous; Bacteria; Bacterial Genome; Bacterial Model; Biological Assay; Breath Tests; Burkholderia; Collection; Communities; Coupled; DNA; Data; Development; Escherichia coli; Gene Cluster; Genes; Genome; Goals; Grant; Growth; Health; Human; Knowledge; Lasso; Libraries; Medicine; Mining; Modeling; Natural Products; Organism; Pathway interactions; Peptide Biosynthesis; Peptides; Play; Privatization; Process; Production; Proteobacteria; Regulation; Regulator Genes; Reporter; Role; Source; Structural Genes; Structure; Taxonomy; Technology; Testing; Transcription Coactivator; base; bioactive natural products; drug discovery; genome sequencing; interest; mass cell culture; novel; peptide B; promoter; resistance gene; synthetic biology; tool; transcriptome sequencing

Project Summary Molecules isolated from living systems have consistently served as leads in drug discovery. However, two of the main challenges of drug discovery from natural products are the high rate of rediscovery of known compounds using traditional technologies and low yields. Recent advances in genome sequencing and synthetic biology have spurred a renewed interest in natural product discovery in the private and academic sectors. For instance, genome mining of underexplored taxa increases the chances of novel compound discovery. Moreover, robust host organisms can facilitate discovery efforts by overcoming the common low-yield hurdle. Bacteria belonging to the Burkholderiales order of -Proteobacteria are an emerging source of natural products. We have previously achieved high yield production of autologous polyketide-nonribosomal peptide spliceostatins in a Burkholderia species. We propose to understand, develop, and apply this strain as a host to discover and produce natural products from Burkholderiales and potentially other -Proteobacteria. Although Escherichia coli has long been used as a model bacterial host, the synthetic biology community is moving away from the idea that “one host fits all” to instead have hosts tailored to the source of biosynthetic gene clusters. We hypothesize that the Burkholderia sp. in question can be used as a host to streamline the discovery process by producing heterologous natural products in high yields. Our preliminary data supports this hypothesis as heterologous expression of a model gene cluster encoding the lasso peptide capistruin in this strain led to capistruin production in yields that are at least 65-fold, and up to 580-fold higher than with E. coli. In Aim 1 we propose to test/understand the host by a) testing the breath of the host in terms of source genera within the Burkholderiales and other -Proteobacteria while discovering lasso peptides; and b) investigating the regulation of autologous spliceostatin biosynthesis with the ultimate goal of deriving regulatory parts for heterologous pathway construction. In Aim 2, we will develop the host as a tool for natural product discovery by discovering and characterizing a promoter library and by generating a minimized genome. In Aim 3 we will apply the host by discovering natural products from a newly built Burkholderiales environmental collection. This project is expected to provide tools that will streamline and accelerate natural product discovery from a promising yet underexplored source. The tools and knowledge obtained will be applied to discover novel, bioactive natural products via genome mining of a Burkholderiales strain collection.

项目概要 从生命系统中分离出来的分子一直是药物发现的先导者。然而，其中的两个 从天然产物中发现药物的主要挑战是已知化合物的高重新发现率 使用传统技术且产量低。基因组测序和合成生物学的最新进展 激发了私营和学术部门对天然产物发现的新兴趣。例如， 对尚未探索的类群进行基因组挖掘增加了发现新化合物的机会。此外，坚固耐用 宿主生物可以通过克服常见的低产障碍来促进发现工作。细菌归属 -Proteobacteria 的伯克霍尔德氏菌目是天然产物的新兴来源。我们之前有过 在伯克霍尔德杆菌中实现了自体聚酮化合物-非核糖体肽剪接抑素的高产率生产 物种。我们建议理解、开发和应用这种菌株作为宿主来发现和生产天然的 来自伯克霍尔德氏菌目和潜在的其他 -变形菌门的产品。尽管大肠杆菌长期以来 作为模型细菌宿主，合成生物学界正在摆脱“一个宿主适合一个宿主”的想法。 all”来代替针对生物合成基因簇的来源定制的宿主。我们假设 伯克霍尔德氏菌有问题的可以用作主机，通过生成来简化发现过程 异源天然产物产量高。我们的初步数据支持这一假设是异源的 编码套索肽 Capistruin 的模型基因簇在该菌株中的表达导致 Capistruin 的产生 产量比大肠杆菌高至少 65 倍、最多 580 倍。在目标 1 中，我们建议 通过 a) 测试宿主在伯克霍尔德氏菌目中的源属方面的呼吸来测试/了解宿主 和其他 -变形菌，同时发现套索肽； b) 调查自体移植的监管 剪接抑素生物合成，最终目标是衍生异源途径的调节部分 建造。在目标 2 中，我们将通过发现和开发宿主作为天然产物发现的工具。 表征启动子文库并生成最小化的基因组。在目标 3 中，我们将通过以下方式应用主机 从新建的伯克霍尔德氏菌环境收藏中发现天然产品。该项目预计 提供工具来简化和加速从有前途但尚未充分探索的天然产物的发现 来源。获得的工具和知识将用于发现新颖的、具有生物活性的天然产品 伯克霍尔德氏菌菌株库的基因组挖掘。