Secondary metabolites are an extremely diverse and important group of natural products with industrial and biomedical implications. Advances in metabolic engineering of both native and heterologous secondary metabolite producing organisms have allowed the directed synthesis of desired novel products by exploiting their biosynthetic potentials. Metabolic engineering utilises knowledge of cellular metabolism to alter biosynthetic pathways. An important technique that combines chemical synthesis with metabolic engineering is mutasynthesis (mutational biosynthesis; MBS), which advanced from precursor-directed biosynthesis (PDB). Both techniques are based on the cellular uptake of modified biosynthetic intermediates and their incorporation into complex secondary metabolites. Mutasynthesis utilises genetically engineered organisms in conjunction with feeding of chemically modified intermediates. From a synthetic chemist's point of view the concept of mutasynthesis is highly attractive, as the method combines chemical expertise with Nature's synthetic machinery and thus can be exploited to rapidly create small libraries of secondary metabolites. However, in each case, the method has to be critically compared with semi- and total synthesis in terms of practicability and efficiency. Recent developments in metabolic engineering promise to further broaden the scope of outsourcing chemically demanding steps to biological systems.

The timing of introduction of the unusually placed Delta(11,13) diene system in ansamitocin (AP) biosynthesis was probed by synthesizing optically active potential tri- and tetraketide intermediates as their SNAC thioesters. An AP-nonproducing mutant Actinosynnema pretiosum was complemented by the R enantiomer of the triketide and by the tetraketide with rearranged double bonds, but not by the tetraketide carrying the double bonds in conjugation to the thioester function. The results show that the double bonds are installed in their final positions during processing of the nascent polyketide on module 3 of the asm PKS and that KS4 of the PKS acts as a gatekeeper which accepts only a tetraketide with shifted double bonds as substrate for further processing.

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