Studies on terpenoids produced by actinomycetes-Screening for new bioactive compounds

本项目旨在从放线菌中分离新的萜类化合物，并研究其化学和生物学特性。为了达到这一目的，我们首先尝试用甲基戊酸途径的关键酶HMG-CoA还原酶基因作为PCR探针，分离出具有甲基戊酸途径的放线菌，然后从微生物中分离出萜类。与我们的预期相反，分离出的阳性菌株数量很少，但我们成功地纯化了几个新的萜类化合物。其中一种化合物被证实是一种新的二萜衍生物，具有异松二烯骨架，从未作为放线菌代谢产物被分离出来。由于专利申请，该化合物的详细性质尚未公开，但其对EGF蛋白酪氨酸激酶具有较强的抑制活性，其详细的生物学活性正在研究中。从另一株链霉菌中也分离到了新的萜类化合物。为了揭示甲羟戊酸途径与萜类生物合成之间的关系，本项目对产萜类放线菌的相关基因进行了研究。结果表明，甲羟戊酸途径基因簇周围存在萜类生物合成相关基因。这一发现表明甲羟戊酸途径与萜类化合物的产生密切相关。这些基因的表达是在生产生物生长的后期，当次生代谢物开始生产时观察到的。用甲羟戊酸途径抑制一株链霉菌的甲羟戊酸途径可引起一种新化合物mevashuntin的积累。所获得的信息预计将对放线菌生产萜类化合物的研究具有重要意义。

The purpose of this project is to isolate new terpenoids from Actinomycetes and to investigate their chemical and biological properties. In order to achieve this purpose, we first attempted to isolate actinomycetes possessing the mevalonate pathway by using the HMG-CoA reductase gene as a probe for PCR, the key enzyme of the mevalonate pathway, and then to isolate terpenoids from the microorganisms. Contrary to our expectation, the number of positive strains isolated was very small, but we succeeded in purification of several new terpenoids. Among them, one compound proved to be a new diterpene derivative with the isopimaradiene skeleton that has never been isolated as actinomycetes metabolites. Although detail properties of this new compound can not be disclosed due to patent application, it showed strong inhibitory activity to EGF protein tyrosine kinase and its detailed biological activities are now under investigation. Another new terpenoid was also isolated from another strain of Streptomyces.In order to disclose the relationship between the mevalonate pathway and terpenoid biosynthesis, the relevant genes of actinomycetes producing terpenoids were investigated in this project. As a result, it turned out that genes related to terpenoid biosynthesis existed around the mevalonate pathway gene cluster. This finding shows that the mevalonate pathway is strictly linked to production of terpenoids. Expression of these gene was observed at the later stage of the growth of the producing organisms when the production of secondary metabolites started.Inhibition of the mevalonate pathway in one Streptomyces strain with the mevalonate pathway caused accumulation of a new compound, mevashuntin. The obtained information is expected to be important for studies related to production of terpenoids by actinomycetes.