Bacillus subtilis Utilizes Decarboxylated S-Adenosylmethionine for the Biosynthesis of Tandem Aminopropylated Microcin C, a Potent Inhibitor of Bacterial Aspartyl-tRNA Synthetase

Abstract

The biosynthetic pathways of natural products involve unusual biochemical reactions catalyzed by unique enzymes. Aminopropylation, although apparently simple, is an extremely rare modification outside polyamine biosynthesis. The canonical pathway used in the biosynthesis of peptide-adenylate antibiotic microcin C of E. coli (Eco-McC) entails alkylation by the S-adenosyl-methionine-derived 3-amino-3-carboxypropyl group of the adenylate moiety and subsequent decarboxylation to yield the bioactive aminopropylated compound. Here, we report the structure and biosynthesis of a new member of the microcin C family of antibiotics, Bsu-McC, produced by Bacillus subtilis MG27, which employs an alternative aminopropylation pathway. Like Eco-McC, Bsu-McC consists of a peptide moiety that facilitates prodrug import into susceptible bacteria and a warhead, a nonhydrolyzable modified isoasparaginyl-adenylate, which, when released into the cytoplasm, binds aspartyl-tRNA synthetase (AspRS) inhibiting translation. In contrast to the Eco-McC, whose warhead carries a single aminopropyl group attached to the phosphate moiety of isoasparaginyl-adenylate, the warhead of Bsu-McC is decorated with a tandem of two aminopropyl groups. Our in silico docking of the Bsu-McC warhead to the AspRS-tRNA complex suggests that two aminopropyl groups form extended interactions with the enzyme and tRNA, stabilizing the enzyme–inhibitor complex. We show that tandem aminopropylation results in a 32-fold increase in the biological activity of peptidyl-adenylate. We also show that B. subtilis adopted an alternative pathway for aminopropylation in which two homologous 3-aminopropyltransferases utilize decarboxylated S-adenosylmethionine as a substrate. Additionally, Bsu-McC biosynthesis alters the social behavior of the B. subtilis producer strain, resulting in a sharp decrease in their ability to form biofilms.