Abstract：
Triterpenoids are a type of specialized metabolites that exhibit a wide range of biological activities. However, the availability of some minor triterpenoids in nature is limited, which has hindered our understanding of their pharmacological potential. To overcome this limitation, heterologous biosynthesis of triterpenoids in yeast has emerged as a promising and time-efficient production platform for obtaining these minor compounds. In this study, we analyzed the transcriptomic data of Enkianthus chinensis to identify one oxidosqualene cyclase (EcOSC) gene and four CYP716s. Through heterologous expression of these genes in yeast, nine natural pentacyclic triterpenoids, including three skeleton products (1-3) produced by one multifunctional OSC and six minor oxidation products (4-9) catalyzed by CYP716s, were obtained. Of note, we discovered that CYP716E60 could oxidize ursane-type and oleanane-type triterpenoids to produce 6β-OH derivatives, marking the first confirmed C-6β hydroxylation in an ursuane-type triterpenoid. Compound 9 showed moderate inhibitory activity against NO production and dose-dependently reduced IL-1β and IL-6 production at the transcriptional and protein levels. Compounds 1, 2, 8, and 9 exhibited moderate hepatoprotective activity with the survival rates of HepG2 cells from 61% to 68% at 10 μM.
摘要：
三萜类化合物是一类具有广泛生物活性的特殊代谢产物。然而,自然界中一些次要三萜类化合物的可用性有限，这阻碍了我们对它们药理潜力的理解。为了克服这个限制,酵母中三萜类化合物的异源生物合成已成为获得这些次要化合物的有前途且省时的生产平台。在这项研究中,我们分析了Enkianthus的转录组数据，以鉴定一个氧化角鲨烯环化酶（EcOSC）基因和四个CYP716s。通过这些基因在酵母中的异源表达，九种天然五环三萜类化合物，包括由一种多功能OSC产生的三种骨架产物（1-3）和由CYP716s催化的六种次要氧化产物（4-9），已获得。值得注意的是，我们发现CYP716E60可以氧化ursane型和齐墩烷型三萜类化合物产生6β-OH衍生物，标志着乌苏烷型三萜类化合物中首次确认的C-6β羟基化。化合物9对NO产生显示中等抑制活性,并在转录和蛋白质水平上剂量依赖性地减少IL-1β和IL-6的产生。化合物1、2、8和9表现出中等的肝保护活性,在10μM时HepG2细胞的存活率为61%至68%。
