Genome Mining of Fungal Aza-Polycyclic Natural Products Derived from Arginine-Containing Cyclodipeptides

Arginine-containing cyclodipeptide synthases (RCDPSs) from fungi constitute a new family of tRNA-dependent enzymes that can biosynthesize cyclo-Arg-Xaa dipeptides. The incorporation of an arginine residue significantly expands the chemical space of guanidine-containing natural products. Here, we mined fungal biosynthetic gene clusters (BGCs) containing different RCDPS to discover aza-polycyclic natural products. The pno BGC from Aspergillus pseudonomius produced pentacyclic pyrroloindoline diketopiperazines (DKPs) of which the arginine side chain is oxidatively cyclized into a guanidino-proline. Two RCDPS-encoding BGCs, car and esh from Aspergillus carlsbadensis and Eupenicillium shearii, respectively, produced DKPs connected to five- and seven-membered spirocycles as a result of oxidative cyclization of the guanidino group catalyzed by α-ketoglutarate/Fe(II)-dependent oxygenases. The esh pathway involves a tandem cyclization and epimerization to generate the guanidino-bridged tricyclo-[3.3^1,2.2.2]-piperazinedione core. The aza-polycyclic structures characterized in this work demonstrate the potential of using RCDPS as a starting point for the discovery of new natural products.