STTR Phase I: Bio-based Manufacturing of An Anticoagulant Precursor 4-Hydroxycoumarin

STTR 第一阶段：抗凝血前体 4-羟基香豆素的生物基制造

• 批准号: 1549724
• 负责人: Yuheng Lin
• 金额: $ 22.5万
• 依托单位: BiotecEra Inc.
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-01-01 至 2017-06-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1549724&HistoricalAwards=false
• 关键词: STTR; Phase; Bio; based; Manufacturing

The broader impact/commercial potential of this Small Business Technology Transfer (STTR) project is to establish a bio-based manufacturing process for the production of the commodity chemical 4-hydroxycoumarin (4-HC) in an economical and renewable way. 4-HC is a direct synthetic precursor used to manufacture widely used oral anticoagulants, such as warfarin, acenocoumarol, and phenprocoumon. In addition, 4-HC-derived anticoagulants are commonly used as rodenticides to kill rodents. Traditionally, 4-HC is commercially manufactured through chemical synthesis using petro-related aromatic chemicals as starting materials, which is neither economical nor environmentally friendly. The proposed technology to be developed in this project is expected to dramatically lower the production cost, and reduce the reliance on petro chemicals. This STTR Phase I project proposes to create an optimal 4-HC-producing microbial strain that can be readily used for large-scale production. The strain is expected to have high genetic stability without the need for antibiotics or an inducer for production. Preliminary research has shown the microbial production of 4-HC from a renewable carbon source, glycerol, in shake flasks, which demonstrates great commercialization potential. However, the approach at this stage still uses host-plasmid systems that involve antibiotics and the inducer IPTG. This will not only increase the production cost but also will raise environmental concerns. Additionally, host-plasmid systems may cause instability in genetic properties, which is undesirable for large-scale production of commodity chemicals. To address these issues and achieve the goal of this STTR Phase I project, the following research objectives will be pursued: Integration of constitutively expressed 4-HC biosynthetic pathway genes into E. coli chromosomal DNA and its optimization; and inactivation of the E. coli endogenous enzymes responsible for unwanted degradation of 4-HC biosynthetic intermediates.

该小企业技术转让 (STTR) 项目更广泛的影响/商业潜力是建立一种生物基制造工艺，以经济且可再生的方式生产商品化学品 4-羟基香豆素 (4-HC)。 4-HC 是一种直接合成前体，用于制造广泛使用的口服抗凝剂，例如华法林、醋硝香豆素和苯丙香豆素。此外，4-HC衍生的抗凝剂通常用作杀鼠剂来杀死啮齿动物。传统上，4-HC的商业生产是采用石油相关芳烃化学品为原料，通过化学合成的方式进行的，既不经济也不环保。该项目拟开发的技术预计将大幅降低生产成本，并减少对石化产品的依赖。该 STTR 第一阶段项目旨在创建一种可轻松用于大规模生产的最佳 4-HC 生产微生物菌株。该菌株预计具有较高的遗传稳定性，无需抗生素或诱导剂即可生产。初步研究表明，在摇瓶中利用可再生碳源甘油微生物生产 4-HC，具有巨大的商业化潜力。然而，现阶段的方法仍然使用涉及抗生素和诱导剂 IPTG 的宿主质粒系统。这不仅会增加生产成本，还会引起环境问题。此外，宿主-质粒系统可能导致遗传特性不稳定，这对于大规模生产商品化学品来说是不希望的。为了解决这些问题并实现 STTR 一期项目的目标，将追求以下研究目标： 将组成型表达的 4-HC 生物合成途径基因整合到大肠杆菌染色体 DNA 中并对其进行优化；导致 4-HC 生物合成中间体降解的大肠杆菌内源酶失活。