The discovery of antibiotics for the treatment of bacterial infections has been one of the most important medical achievements of the past 60 years.[1] However, the emergence of bacterial resistance to current antibiotics is a major challenge faced by the medical community. The emergence of resistance is inevitable, and hence the continued discovery of novel chemotype antibiotics with novel modes of action is critical to overcome drug-resistant organisms.[2] Several drugdiscovery strategies have emerged, including incremental improvements to existing antibiotics by chemical manipulation and the search for novel drug targets on the basis of genomic approaches. An alternative strategy is to study the biochemical pathways or processes inhibited by known antibiotics and look for as yet unexploited targets within a pathway. A major benefit of employing such an approach is that these pathways are validated targets for drug intervention. The fatty acid synthesis is one such pathway. The targets we chose to study were the fatty acid biosynthesis condensing enzymes FabH and FabF, which are conserved and essential for bacterial viability.[3–6] The FabF target has been the subject of studies for over 20 years, and two classes of inhibitors have been reported:[7–12] cerulenin [13] and thiolactomycin [14–16] and its analogues. Cerulenin is a covalent modifier and a selective inhibitor of FabF, whereas thiolactomycin is a dual inhibitor of FabH and FabF. However, both exhibit poor antibacterial activity (Staphylococcus aureus MIC (minimum inhibitory concentration) 64 μg mLÀ1).[17] Potent inhibitors of these enzymes are expected to allow the development of antibiotics with no cross-resistance to existing drugs. We employed a differential-sensitivity method in which each target was differentially expressed by using antisense methodology under control of a xylose-inducible promoter.[18]

In this method, the strain expressing fabF/fabH antisense RNA exhibits hypersensitivity to FabH-and/or FabF-specific inhibitors relative to a control strain.[18] This allowed us to develop a target-based whole-cell high-throughput-screening assay for the discovery of FabH and FabF inhibitors.[19, 20] Recent screening of natural product extracts by employing this differential-sensitivity whole-cell two-plate agardiffusion assay [19] led us to discover platensimycin (1) as a