Engineering Terpene Synthases for Increased Productivity

工程萜烯合成以提高生产率

• 批准号: 7113465
• 负责人: Jennifer Rachel Anthony
• 金额: $ 4.04万
• 依托单位: UNIVERSITY OF CALIFORNIA BERKELEY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-05-01 至 2007-04-13
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7113465
• 关键词: Escherichia coli; alkyltransferase; bacterial genetics; bacterial proteins; bioreactors; biotechnology; directed evolution; enzyme activity; microorganism mass culture; microorganism metabolism; postdoctoral investigator; protein engineering; solubility; synthetic enzyme; terpenes; transfection

DESCRIPTION (provided by applicant): Terpenes are the most diverse class of natural products, with 30,000 known members. A number of characterized terpenes form commercially useful products, including the anticancer agent Taxol. Large- scale production of terpenes is difficult because current techniques rely on complex chemical synthesis or inefficient extraction from biological sources. To address the demand for therapeutic terpenes the Keasling laboratory has engineered Escherichia coli to produce amorpha-4,11-diene, the precursor for the anti- malarial compound, artemisinin. However, adequate expression of the terpene synthase, amorphadiene synthase (ADS), has become a major obstacle to increased amorphadiene production. Terpene synthases catalyze the cyclization of polyprenyl diphosphates to form a terpene backbone, which is the first committed step in all terpene biosynthesis. In this proposal, we outline an approach to improve terpene synthase expression in E. coli by increasing both solubility and activity,of the model terpene synthase, ADS, as well as to optimize flux through this enzyme to increase amorphadiene production. Altered ADS enzymes will be analyzed and a model linking terpene synthase sequence to enzyme function will be established.

描述（由申请人提供）：萜烯是最多样化的天然产品，拥有30,000名已知成员。许多特征的萜烯形成了商业上有用的产品，包括抗癌剂紫杉醇。大规模的萜烯产生很困难，因为当前技术依赖于复杂的化学合成或从生物来源中提取效率低下。为了满足对治疗性萜烯的需求，Keasling实验室已经设计了大肠杆菌以产生氨基氨基甲烷-4,11-二烯，这是抗疟疾化合物的前体Artemisinin，Artemisinin。然而，萜烯合酶，氨基苯二烯合酶（ADS）的充分表达已成为增加氨基化产量增加的主要障碍。萜烯合酶会催化聚丙烯二磷酸酯的环化形成萜烯骨架，这是所有萜烯生物合成的第一步。在该提案中，我们概述了一种通过增加氧化烯合酶ADS的溶解度和活性来改善大肠杆菌中萜烯合酶表达的方法，并通过该酶优化通量以增加非磷灰二烯产生。将分析变化的AD酶，并将建立将萜烯合酶序列与酶功能联系起来的模型。