(160at) Quinolactacin Biosynthesis Involves Nrpss Catalyzed Dieckmann Condensation to Form the Quinolone-?-Lactam Hybrid

Quinolactacins (QUL) are novel fungal alkaloids that feature a quinolone-γ-lactam hybrid, which is a potential pharmacophore for the treatment of cancer and Alzheimer’s disease. The biomimetic synthesis of QUL from tryptamine required eight-step reactions with a low yield of ~8%. Understanding how nature builds such a unique quinolone-γ-lactam pharmacophore in a highly selective and efficient manner can reveal useful enzymatic mechanisms and lead to the discovery of new bioactive molecules. We first set out to identify and characterize the qul biosynthetic gene cluster. Based on bioinformatics and genetics analysis, one cluster encodes two separate single-module NRPSs, QulA and QulB, a methyltransferase QulM, indoleamine-2,3-dioxygenase (QulI), a putative FMN-dependent dehydrogenase (QulF) and a putative monocarboxylate permease (QulP) are found to be responsible for QUL biosynthesis. We further characterized this gene cluster and revealed an unusual β-keto acid (N-methyl-2-aminobenzoylacetate) precursor that is derived from the primary metabolite L-kynurenine via methylation, oxidative decarboxylation, and amide hydrolysis reactions. Interestingly, an amidase located outside of the qul gene cluster was also confirmed involving this pathway which catalyze the amide hydrolysis of amide hydrolysis to produce an unstable β-keto acid precursor. In vitro assays reveal two single-module NRPSs that incorporate the β-keto acid and L-isoleucine, followed by Dieckmann condensation, to form the quinolone-γ-lactam. Notably, the bioconversion from L-kynurenine to the β-keto acid is a unique strategy employed by Nature to decouple R* domain-containing NRPS from PKS machinery, which expands the paradigm for the biosynthesis of quinolone-γ-lactam natural products via Dieckmann condensation.