Genome-based discovery and total synthesis of janustatins, potent cytotoxins from a plant-associated bacterium

Host-associated bacteria are increasingly being recognized as underexplored sources of bioactive natural products with unprecedented chemical scaffolds. A recently identified example is the plant-root-associated marine bacterium Gynuella sunshinyii of the chemically underexplored order Oceanospirillales. Its genome contains at least 22 biosynthetic gene clusters, suggesting a rich and mostly uncharacterized specialized metabolism. Here, in silico chemical prediction of a non-canonical polyketide synthase cluster has led to the discovery of janustatins, structurally unprecedented polyketide alkaloids with potent cytotoxicity that are produced in minute quantities. A combination of MS and two-dimensional NMR experiments, density functional theory calculations of 13 C chemical shifts and semiquantitative interpretation of transverse rotating-frame Overhauser effect spectroscopy data were conducted to determine the relative configuration, which enabled the total synthesis of both enantiomers and assignment of the absolute configuration. Janustatins feature a previously unknown pyridodihydropyranone heterocycle and an unusual biological activity consisting of delayed, synchronized cell death at subnanomolar concentrations. In silico chemical prediction of a polyketide synthase gene cluster in the bacterium Gynuella sunshinyii has led to the discovery of a class of natural products called janustatins. The absolute configuration of the stereocentres in these compounds was determined by a combination of techniques including DFT calculations and 2D NMR experiments—and finally confirmed by total synthesis. Janustatins were found to cause delayed, synchronized cell death at subnanomolar concentrations.