Identification of diterpene biosynthetic gene clusters and functional analysis of labdane-related diterpene cyclases in Phomopsis amygdali
Identification of diterpene biosynthetic gene clusters and functional analysis of labdane-related diterpene cyclases in Phomopsis amygdali
复制标题
DOI：
10.1271/bbb.70790
复制
发表时间：
2008-04-01
影响因子：
1.6
通讯作者：
Sassa, Takeshi
中科院分区：
文献类型：
--
作者：
Toyomasu, Tomonobu;Nuda, Rie;Sassa, Takeshi
利用基因组步移技术，从产丝孢菌素的扁桃拟茎点霉（Phomopsis amygdali）中分离到两个二萜生物合成基因簇，分别为编码香叶基香叶基二磷酸（GGDP）脱氢酶的PaGGS 1和PaGGS 4。二萜环化酶样基因PaDC 1和PaDC 2分别位于PaGGS 1和PaGGS 4附近。这两种酶的氨基酸序列与真菌劳丹相关的二萜环化酶的氨基酸序列相似。重组PaDC 1主要通过（+）-柯巴基二磷酸（CDP）将GGDP转化为叶枝烷-16 α-醇和痕量的几种劳丹烷相关的烃，这些烃已从苦杏仁P. amygdali F6菌丝体中鉴定出来。由于在苦杏仁F6菌丝体中尚未鉴定出phyllocladan-16 α-醇，我们从菌丝体中分离出phyllocladan-16 α-醇。重组PaDC 2将GGDP转化为（+）-CDP。此外，我们还分离出了一种新的二萜类化合物phyllocladan-11 alpha，16 alpha，18-triol，它可能是phyllocladan-16 alpha-ol在菌丝体中的代谢产物。我们建议，基因组步行提供了一个有用的策略，在真菌中发现新的天然产物。
Two diterpene biosynthesis gene clusters in the fusicoccin-producing fungus, Phomopsis amygdali, were identified by genome walking from PaGGS1 and PaGGS4 which encode the geranylgeranyl diphosphate (GGDP) synthases. The diterpene cyclase-like genes, PaDC1 and PaDC2, were respectively located proximal to PaGGS1 and PaGGS4. The amino acid sequences of these two enzymes were similar to those of fungal labdane-related diterpene cyclases. Recombinant PaDC1 converted GGDP mainly into phyllocladan-16 alpha-ol via (+)-copalyl diphosphate (CDP) and trace amounts of several labdane-related hydrocarbons which had been identified from the P. amygdali F6 mycelia. Since phyllocladan-16 alpha-ol had not been identified in P. amygdali F6 mycelia, we isolated phyllocladan-16 alpha-ol from the mycelia. Recombinant PaDC2 converted GGDP into (+)-CDP. Furthermore, we isolated the novel diterpenoid, phyllocladan-11 alpha,16 alpha,18-triol, which is a possible metabolite of phyllocladan-16 alpha-ol in the mycelia. We propose that genome walking offers a useful strategy for the discovery of novel natural products in fungi.