Reconstitution of biosynthetic machinery for indole-diterpene paxilline in Aspergillus oryzae.

Koichi Tagami, Chengwei Liu, Atsushi Minami, Motoyoshi Noike, Tetsuya Isaka, Shuhei Fueki, Yoshihiro Shichijo, Hiroaki Toshima, Katsuya Gomi, Tohru Dairi, Hideaki Oikawa

Division of Chemistry, Graduate School of Science, Hokkaido University, Sapporo 060-0810, Japan.

Journal of the American Chemical Society 2013 Jan 30

filter terms:

Indole-diterpenes represented by paxilline share a common pentacyclic core skeleton derived from indole and geranylgeranyl diphosphate. To shed light on the detailed biosynthetic mechanism of the paspaline-type hexacyclic skeleton, we examined the reconstitution of paxilline biosynthetic machinery in Aspergillus oryzae NSAR1. Stepwise introduction of the six pax genes enabled us to isolate all biosynthetic intermediates and to synthesize paxilline. In vitro and in vivo studies on the key enzymes, prenyltransferase PaxC and cyclase PaxB, allowed us to elucidate actual substrates of these enzymes. Using the isolated and the synthesized epoxide substrates, the highly intriguing stepwide epoxidation/cyclization mechanism for the construction of core structure has been confirmed. In addition, we also demonstrated "tandem transformation" to simultaneously introduce two genes using a single vector (paxG/paxB, pAdeA; paxP/paxQ, pUNA). This may provide further option for the reconstitution strategy to synthesize more complex fungal metabolites.

Citation

Koichi Tagami, Chengwei Liu, Atsushi Minami, Motoyoshi Noike, Tetsuya Isaka, Shuhei Fueki, Yoshihiro Shichijo, Hiroaki Toshima, Katsuya Gomi, Tohru Dairi, Hideaki Oikawa. Reconstitution of biosynthetic machinery for indole-diterpene paxilline in Aspergillus oryzae. Journal of the American Chemical Society. 2013 Jan 30;135(4):1260-3