紫金牛中一个新颖底物杂泛性O-甲基转移酶

A novel O-methyltransferase with substrate promiscuity from Ardisia japonica

O-甲基转移酶(O-methyltransferases,OMTs)是许多天然产物生物合成途径中的关键后修饰酶之一,O-甲基化不仅可降低化合物的化学反应活性,而且能显著改善其溶解性、稳定性等理化性质和生物活性等。本研究根据紫金牛转录组测序结果,首次从紫金牛中克隆获得1条OMT基因(命名为AjOMT1),并成功进行了大肠杆菌异源表达,体外酶催化活性结果表明重组AjOMT1能高效催化槲皮素发生O-甲基化反应。底物谱考察结果表明,AjOMT1能催化黄酮类、茋类、香豆素类、生物碱类、苯丙素类等不同结构类型的化合物进行甲基化反应,具有非常宽泛的底物谱;AjOMT1偏好于具邻二酚羟基的化合物,并表现出位置选择性。本研究结果表明,AjOMT1可用作O-甲基化工具酶实现不同类型化合物的O-甲基化、获取多样化的活性甲基化产物,可为药物发现提供新方法,并可为天然产物合成生物学研究提供普适性元件。

O-methyltransferases(OMTs)are one of the key tailoring enzymes in the biosynthesis of many natural products,O-methylation can not only reduce the reactivity of natural products,but also alter their solubility,stability and biological activities.Based on the transcriptome data of Ardisia japonica,a full-length cDNA sequence of candidate OMT(termed as AjOMT1)was cloned by reverse transcription-polymerase chain reaction(RT-PCR)and expressed in Escherichia coli(E.coli)for the first time.In vitro enzyme catalytic activity assay showed that the recombinant AjOMT1 could effectively catalyze quercetin to form O-methylated products.Most importantly,AjOMT1 showed unprecedented substrate promiscuity towards structurally various compounds including flavonoids,stilbenes,coumarins,alkaloids and phenylpropanoids,especially preferring to the compounds with adjacent phenolic hydroxyl groups,and showed regio-selectivity.These results suggested that AjOMT1 could be used as the tool enzyme to conduct O-methylation of different types of compounds to produce diverse active methylated products,and provide a new method for drug discovery,even universal part for synthetic biology of natural products.