The α,β-epoxyketone proteasome inhibitor TMC-86A was discovered as a previously unreported metabolite of Streptomyces chromofuscus ATCC49982, and the gene cluster responsible for its biosynthesis was identified via genome sequencing. Incorporation experiments with [(13)C-methyl]l-methionine implicated an α-dimethyl-β-keto acid intermediate in the biosynthesis of TMC-86A. Incubation of the chemically synthesized α-dimethyl-β-keto acid with a purified recombinant flavin-dependent enzyme that is conserved in all known pathways for epoxyketone biosynthesis resulted in formation of the corresponding α-methyl-α,β-epoxyketone. This transformation appears to proceed via an unprecedented decarboxylation-dehydrogenation-monooxygenation cascade. The biosynthesis of the TMC-86A warhead is completed by cytochrome P450-mediated hydroxylation of the α-methyl-α,β-epoxyketone.

Fungus-associated Burkholderia gladioli bacteria use a unique ‘dual-priming’ nonribosomal peptide synthetase to assemble icosalide A1.

The complete stereochemical course and substrate selectivity of the enoyl reductase domain from the fungal polyketide synthase squalestatin tetraketide synthase (SQTKS) have been determined.