Biosynthesis of a N-Acetylated Tricyclic Carbazole with Antithrombotic Activity.

Tricyclic carbazoles are significant pharmacophores. Herein, heterologous expression of the carbazole-3,4-quinone (1) biosynthetic pathway in the chassis host Streptomyces albus J1074 yielded a previously chemosynthesized orthoquinone carbazole (2) and three new N-acetylated carbazoles (3-5). Their structures were established by a combination of HR-ESI-MS, NMR, and X-ray crystallographic analysis. Compound 2, the deaminated precursor of 4 and 5, was enzymatically synthesized, indicating the substrate tolerance of the key enzymes in the bacterial tricyclic-carbazole biosynthetic pathway. Mutagenetic analysis revealed an arylamine N-acetyltransferase homologous gene required for the production of compounds 3-5. Bioactivity analysis using the zebrafish model demonstrated that compound 5 has significant antithrombotic activity, potentially by downregulating the genes involved in the platelet activation and coagulation cascade. These findings expand the natural strategies for structural diversification of the tricyclic carbazole alkaloids.