Elucidation of shanorellin biosynthetic pathway and functional analysis of associated enzymes.
Elucidation of shanorellin biosynthetic pathway and functional analysis of associated enzymes.
复制标题
香诺雷林生物合成途径的阐明和相关酶的功能分析。
作者：
Sato;M.;Yamada;H.;Hotta;K.;Watanabe;K.
由于真菌天然产物的生物合成的聚酮酶经常表现出有用的生物活性，确定和理解的PKS生物合成步骤的机制是非常感兴趣的。从球毛壳菌中分离的一种这样的化合物是山诺瑞林，其生物合成基因簇通过过表达转录因子CgsG而被激活。通过基因敲除技术在C.通过globosum和体外生化分析，我们首次确定了山诺菌素生物合成的基因簇和途径。我们还确定，细胞色素P450 CgsB是负责催化CgsA生物合成的芳香族产物中C5甲基的羟基化。随后，含黄素的单加氧酶CgsF影响中间体的脱羧作用，形成醌产物山诺瑞林。虽然这类天然产物表现出显著的生物活性，但在生产生物体中相关基因簇的沉默导致在常规培养条件下这些化合物的痕量水平生产。生物合成基因簇的转录调节因子的过表达通过将可调节的启动子并入天然真菌基因组中来实现。这种方法使我们能够鉴定shanorellin基因簇，并有效地破译其生物合成途径。
Since fungal natural products biosynthesized by polyketide synthases frequently exhibit useful biological activities, identifying and understanding the mechanism of biosynthetic steps taken by PKSs are of great interest. One such compound isolated from Chaetomium globosum is shanorellin, whose biosynthetic gene cluster was activated by overexpressing the transcription factor, CgsG. Through targeted gene knockout in C. globosum and in vitro biochemical analyses, we determined for the first time the gene cluster and the pathway for the biosynthesis of shanorellin. We also identified that cytochrome P450 CgsB is responsible for catalysing hydroxylation of a methyl group at C5 in the aromatic product biosynthesized by CgsA. Subsequently, flavin-containing monooxygenase CgsF affects decarboxylation of the intermediate to form the quinone product, shanorellin. While this class of natural products exhibit notable biological activities, silencing of relevant gene clusters in the producing organisms results in trace-level production of those compounds under conventional culture conditions. Overexpression of the biosynthetic gene cluster's transcription regulator was achieved by incorporating a regulatable promoter into the native fungal genome. This approach allowed us to identify the shanorellin gene cluster and decipher its biosynthetic pathway effectively.