卟啉生物碱具有多种药理活性;然而,我们对它们的生物合成的了解相对有限。先前的研究根据D环的构型和取代基的数量将阿帕芬生物碱分为两类,并提出了每个类别的初步生物合成途径。在这项研究中,我们鉴定了两种特定的细胞色素P450酶(CYP80G6和CYP80Q5),它们对多年生藤本植物Stephaniatettrandra的(S)配置和(R)配置底物具有不同的活性,阐明了这两个阿帕芬生物碱类别的生物合成机制和立体化学特征。此外,我们表征了两种CYP719C酶(CYP719C3和CYP719C4)催化亚甲基二氧基桥的形成,一个重要的药效学组,在A-和D-环上,分别,阿帕芬生物碱。利用这些关键细胞色素P450酶的功能表征,我们重建了发芽酵母(酿酒酵母)中两种类型的阿波芬生物碱的生物合成途径,用于从头生产化合物,例如(R)-glaziovine,(S)-glaziovine,还有magnoflorine.这项研究为阿帕芬生物碱的生物合成提供了关键的见解,并为通过合成生物学生产这些有价值的化合物奠定了基础。
Aporphine alkaloids have diverse pharmacological activities; however, our understanding of their
biosynthesis is relatively limited. Previous studies have classified aporphine alkaloids into two categories based on the configuration and number of substituents of the D-ring and have proposed preliminary biosynthetic pathways for each category. In this study, we identified two specific cytochrome P450 enzymes (CYP80G6 and CYP80Q5) with distinct activities toward (S)-configured and (R)-configured substrates from the herbaceous perennial vine Stephania tetrandra, shedding light on the biosynthetic mechanisms and stereochemical features of these two aporphine alkaloid categories. Additionally, we characterized two CYP719C enzymes (CYP719C3 and CYP719C4) that catalyzed the formation of the methylenedioxy bridge, an essential pharmacophoric group, on the A- and D-rings, respectively, of aporphine alkaloids. Leveraging the functional characterization of these crucial cytochrome P450 enzymes, we reconstructed the biosynthetic pathways for the two types of aporphine alkaloids in budding yeast (Saccharomyces cerevisiae) for the de novo production of compounds such as (R)-glaziovine, (S)-glaziovine, and magnoflorine. This study provides key insight into the
biosynthesis of aporphine alkaloids and lays a foundation for producing these valuable compounds through synthetic biology.