The Insect Pathogen <em>Serratia marcescens</em> Db10 Uses a Hybrid Non-Ribosomal Peptide Synthetase-Polyketide Synthase to Produce the Antibiotic Althiomycin

<div><p>There is a continuing need to discover new bioactive natural products, such as antibiotics, in genetically-amenable micro-organisms. We observed that the enteric insect pathogen, <em>Serratia marcescens</em> Db10, produced a diffusible compound that inhibited the growth of <em>Bacillis subtilis</em> and <em>Staphyloccocus aureus</em>. Mapping the genetic locus required for this activity revealed a putative natural product biosynthetic gene cluster, further defined to a six-gene operon named <em>alb1–alb6</em>. Bioinformatic analysis of the proteins encoded by <em>alb1–6</em> predicted a hybrid non-ribosomal peptide synthetase-polyketide synthase (NRPS-PKS) assembly line (Alb4/5/6), tailoring enzymes (Alb2/3) and an export/resistance protein (Alb1), and suggested that the machinery assembled althiomycin or a related molecule. Althiomycin is a ribosome-inhibiting antibiotic whose biosynthetic machinery had been elusive for decades. Chromatographic and spectroscopic analyses confirmed that wild type <em>S. marcescens</em> produced althiomycin and that production was eliminated on disruption of the <em>alb</em> gene cluster. Construction of mutants with in-frame deletions of specific <em>alb</em> genes demonstrated that Alb2–Alb5 were essential for althiomycin production, whereas Alb6 was required for maximal production of the antibiotic. A phosphopantetheinyl transferase enzyme required for althiomycin biosynthesis was also identified. Expression of Alb1, a predicted major facilitator superfamily efflux pump, conferred althiomycin resistance on another, sensitive, strain of <em>S. marcescens</em>. This is the first report of althiomycin production outside of the <em>Myxobacteria</em> or <em>Streptomyces</em> and paves the way for future exploitation of the biosynthetic machinery, since <em>S. marcescens</em> represents a convenient and tractable producing organism.</p>