Fungal bicyclo[2.2.2]diazaoctane indole alkaloids represent an important family of natural products with a wide spectrum of biological activities. Although biomimetic total syntheses of representative compounds have been reported, the details of their biogenesis, especially the mechanisms for the assembly of diastereomerically distinct and enantiomerically antipodal metabolites, have remained largely uncharacterized. Brevianamide A represents a basic form of the subfamily bearing a dioxopiperazine core and a rare 3-spiro-ψ-indoxyl skeleton. In this study, we have identified the brevianamide A biosynthetic gene cluster from Penicillium brevicompactum NRRL 864 and elucidated the metabolic pathway. BvnE was revealed to be an essential isomerase/semipinacolase that specifies the selective production of the natural product. Structural elucidation, molecular modelling and mutational analysis of BvnE as well as quantum chemical calculations have provided mechanistic insights into the diastereoselective formation of the 3-spiro-ψ-indoxyl moiety in brevianamide A. This occurs through a BvnE-controlled semipinacol rearrangement and a subsequent spontaneous intramolecular [4+2] hetero-Diels–Alder cycloaddition.

真菌双环[2.2.2]重氮辛烷吲哚生物碱代表着重要的天然产物家族，具有广泛的生物活性。尽管已经报道了代表性化合物的仿生全合成，但是其生物发生的细节，尤其是非对映异构体和对映异构体对映体代谢产物的组装机理，仍未得到充分表征。布雷维亚酰胺A代表带有二氧哌哌嗪核心和罕见的3-螺-ψ-吲哚基骨架的亚家族的基本形式。在这项研究中，我们从短小青霉菌中鉴定了brevianamide A生物合成基因簇NRRL 864阐明了代谢途径。揭示了BvnE是必需的异构酶/半乳糖苷酶，它指定了天然产物的选择性生产。BvnE的结构阐明，分子建模和突变分析以及量子化学计算，为brevianamide A中3-spiro-ψ-indoxyl部分的非对映选择性形成提供了机械学见解。这是通过BvnE控制的半松果酚重排以及随后的反应而发生的自发的分子内[4 + 2]杂Diels-Alder环加成反应。