Identification of a Pyrrole Intermediate Which Undergoes C‐Glycosidation and Autoxidation to Yield the Final Product in Showdomycin Biosynthesis

Showdomycin is a C‐nucleoside bearing an electrophilic maleimide base. Herein, the biosynthetic pathway of showdomycin is presented. The initial stages of the pathway involve non‐ribosomal peptide synthetase (NRPS) mediated assembly of a 2‐amino‐1H‐pyrrole‐5‐carboxylic acid intermediate. This intermediate is prone to air oxidation whereupon it undergoes oxidative decarboxylation to yield an imine of maleimide, which in turn yields the maleimide upon acidification. It is also shown that this pyrrole intermediate serves as the substrate for the C‐glycosidase SdmA in the pathway. After coupling with ribose 5‐phosphate, the resulting C‐nucleoside undergoes a similar sequence of oxidation, decarboxylation and deamination to afford showdomcyin after exposure to air. These results suggest that showdomycin could be an artifact due to aerobic isolation; however, the autoxidation may also serve to convert an otherwise inert product of the biosynthetic pathway to an electrophilic C‐nucleotide thereby endowing showdomycin with its observed bioactivities. C‐nucleoside biosynthesis: The biosynthetic pathway of the C‐nucleoside showdomycin was reconstituted in vitro. Key steps include cyclization of l‐glutamine to yield an oxygen‐labile pyrrole intermediate that serves as the substrate for C‐glycosidation catalyzed by SdmA. Autoxidative maturation of the pyrrole nucleobase to the maleimide moiety of showdomycin can then proceed non‐enzymatically.