Excellence in Research: Biosynthetic investigation of manzamine class alkaloids

卓越研究：曼扎明类生物碱的生物合成研究

• 批准号: 2302454
• 负责人: Madan Kharel
• 金额: $ 46.03万
• 依托单位: University of Maryland Eastern Shore
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-07-01 至 2026-06-30
• 项目状态: 未结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2302454&HistoricalAwards=false
• 关键词: Excellence; Research; Biosynthetic; investigation; manzamine

Manzamines are members of marine alkaloid natural products. They possess complex structures and exhibit a number of biological activities, including anticancer and antibacterial activities. Lack of knowledge of the molecular and biochemical basis for the production of these alkaloids has been a major barrier to the bioengineering of new alkaloid derivatives. This project will provide such knowledge and will serve as the foundation for bioengineering of new manzamine derivatives with potential applications in medicine, agriculture and biotechnology. This project will also provide extensive research experiences for University of Maryland Eastern Shore undergraduate and graduate students, many of whom are from groups underrepresented in STEM, and thus the work will contribute to development of a diverse STEM workforce. Manzamines represent a structurally distinct group of marine alkaloid natural products with little resemblance to other alkaloids. The structural uniqueness of manzamines, including manzamine A arises from the assembly of a complex pentacyclic core attached to a beta-carboline unit. Although over a hundred manzamines that exhibit a wide range of biological activities have been isolated primarily from marine sponges, how organisms assemble building blocks and intermediates to furnish complex structural scaffolds of manzamines has remained a mystery. The main objective of the proposed project is to identify biosynthetic building blocks and pathway intermediates and elucidate key biosynthetic steps involved in the production of manzamine A. The proposed project utilizes a multi-pronged experimental approach including: (A) isotope incorporation experiments to identify biosynthetic building blocks of the beta-carboline and the pentacyclic ring, (B) experiments to identify the enzyme involved in the attachment of these two structural units, (C) complete genome sequence analysis of Micromonospora sp. M42 for identifying manzamine biosynthetic locus, and (D) heterologous production and characterization of select enzymes involved in the manzamine A biosynthetic pathway. The results garnered through this project will pave the way for the discovery of other manzamines through bioinformatic analysis. Manzamine A biosynthetic enzymes with potentially new catalytic activities will be an invaluable addition to the existing repertoire of tools for the further advancement of research projects concerning biosynthetic pathway engineering/synthetic biology, particularly for the production of new manzamine analogs. The proposed project thus offers a biological alternative to the synthetic production of new manzamine alkaloids.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

曼扎胺是海洋生物碱天然产物的成员。它们具有复杂的结构并表现出多种生物活性，包括抗癌和抗菌活性。缺乏对这些生物碱生产的分子和生化基础的了解一直是新生物碱衍生物生物工程的主要障碍。该项目将提供此类知识，并将作为新型曼扎胺衍生物生物工程的基础，在医学、农业和生物技术方面具有潜在应用。该项目还将为马里兰大学东岸分校的本科生和研究生提供丰富的研究经验，其中许多学生来自 STEM 领域代表性不足的群体，因此这项工作将有助于培养多元化的 STEM 劳动力队伍。曼扎胺代表一组结构独特的海洋生物碱天然产物，与其他生物碱几乎没有相似之处。曼扎胺（包括曼扎胺 A）的结构独特性源于连接至 β-咔啉单元的复杂五环核心的组装。尽管主要从海绵中分离出一百多种表现出广泛生物活性的曼扎胺，但生物体如何组装构建单元和中间体以提供复杂的曼扎胺结构支架仍然是一个谜。拟议项目的主要目标是确定生物合成构件和途径中间体，并阐明生产曼扎明 A 所涉及的关键生物合成步骤。拟议项目采用多管齐下的实验方法，包括：(A) 同位素掺入实验来鉴定生物合成β-咔啉和五环环的构建模块，(B) 鉴定参与这两个结构单元连接的酶的实验，(C) 完整的基因组序列分析小单孢菌属 sp. M42 用于识别曼扎明生物合成位点，以及 (D) 参与曼扎明 A 生物合成途径的选定酶的异源生产和表征。通过该项目获得的结果将为通过生物信息分析发现其他曼扎胺铺平道路。具有潜在新催化活性的曼扎明 A 生物合成酶将是对现有工具库的宝贵补充，可进一步推进有关生物合成途径工程/合成生物学的研究项目，特别是新曼扎明类似物的生产。因此，拟议的项目为新曼扎明生物碱的合成生产提供了一种生物替代方案。该奖项反映了 NSF 的法定使命，并通过使用基金会的智力价值和更广泛的影响审查标准进行评估，被认为值得支持。