Heterologous Production of Eleutherobin Precursors

描述（由申请人提供）：萜类化合物是一类具有同样多样和有效生物活性的天然化合物。由于萜类化合物的实际应用取决于提取或合成这些化合物的能力，因此只有最丰富或最容易合成的化合物才被商业开发。这个小企业技术转让第一阶段项目的长期目标是通过经济地生产稀缺的萜烯来促进新癌症化合物的临床测试和使用。刺五加苷和珊瑚素A和B是细胞毒性萜烯，其作用方式类似于非常成功的抗癌药物紫杉醇。然而，供应的限制阻碍了这些令人兴奋的化合物的发展。这项第一阶段的研究将使用创新的方法来识别，克隆和表达负责形成五加素碳骨架的萜烯环化酶。通过检测环化酶的活性，我们将分离并部分测序eunicellane环化酶。然后将这些部分序列用于设计PCR探针以克隆全长环化酶基因。环化酶的序列将作为发现完整生物合成途径的起点。由于海洋动物产生的许多化合物是微生物共生体的产物，环化酶基因允许鉴定与我们感兴趣的化合物相关的生物合成基因簇。在细菌中发现和表达所鉴定的过度产生萜烯前体的生物合成途径将允许在大规模发酵中产生有价值的萜烯。这项研究与加州大学合作进行，将有助于扩大萜烯环化酶的结构，功能和进化领域，并将为研究生和本科生提供教育机会。这项技术可导致开发出有前途的新的以铽为基础的药物，其生产成本对消费者和环境都较低，因为这会破坏自然资源和释放合成化学废液。

DESCRIPTION (provided by applicant): Terpenoids are a diverse class of natural compounds with equally diverse and potent bioactivities. Because the practical use of terpenoids hinges on the ability to extract or synthesize these compounds, only the most abundant or easily synthesized compounds have been commercially developed. The long-term goal of this Small Business Technology Transfer Phase I project is to facilitate the clinical testing and use of new cancer compounds through the economical production of otherwise scarce terpenes. Eleutherobin and sarcodictyin A and B are cytotoxic terpenes that act in a manner analogous to very successful anti-cancer drug Taxol. Supply limitations, however, has precluded the development of these exciting compounds. This Phase I research will use innovative methods to identify, clone and express the terpene cyclase responsible for the formation of the eleutherobin carbon backbone. By detecting the activity of the cyclase, we will isolate and partially sequence the eunicellane cyclase. These partial sequences will then be used to design PCR probes to clone the full length cyclase gene. The sequence of the cyclase will act as a starting point in the discovery of the full biosynthetic pathway. As many compound produced by marine animals are the product of microbial symbionts, the cyclase gene allow the identification of biosynthetic gene clusters associated with our compound of interest. The discovery and expression of the identified biosynthetic pathway in bacteria that over-produce terpene precursors will allow the production of valuable terpenes in large scale fermentations. This research, conducted in collaboration with the University of California, will contribute to the expanding field of terpene cyclase structure, function and evolution and will provide educational opportunities for graduate and under-graduate students. This technology could result in the development of promising new terpene-based drugs produced at a lower cost to consumers and to the environment, in terms of destruction of natural resources and the release of synthetic chemical effluents.