Class II terpene cyclases, such as oxidosqualene and squalene-hopene cyclases, catalyze some of the most complex polycyclization reactions. They minimally exhibit a β,γ-didomain architecture that has been evolutionarily repurposed in a wide range of terpene-processing enzymes and likely resulted from a fusion of unidentified monodomain proteins. Although single domain class I terpene cyclases have already been identified, single domain class II terpene cyclases have not been previously reported. Here we report high-resolution X-ray structures of a monodomain class II cyclase, merosterolic acid synthase (MstE). With a minimalistic β-domain architecture, this cyanobacterial enzyme is able to construct four rings in cytotoxic meroterpenoids with a sterol-like topology. The structures with bound substrate, product, and inhibitor provide detailed snapshots of a cyclization mechanism largely governed by residues located in a noncanonical enzyme region. Our results complement the few known class II cyclase crystal structures, while also indicating that archaic monodomain cyclases might have already catalyzed complex reaction cascades.

Bacterial multimodular polyketide synthases (PKSs) are large enzymatic assembly lines that synthesize many bioactive natural products of therapeutic relevance. While PKS catalysis is mostly based on fatty acid biosynthetic principles, polyketides can be further diversified by post‐PKS enzymes. Here, we characterized a remarkably versatile trans‐acyltransferase (trans‐AT) PKS from Serratia that builds structurally complex macrolides via more than ten functionally distinct PKS modules. In the oocydin PKS, we identified a new oxygenation module that α‐hydroxylates polyketide intermediates, a halogenating module catalyzing backbone γ‐chlorination, and modular O‐acetylation by a thioesterase‐like domain. These results from a single biosynthetic assembly line highlight the expansive biochemical repertoire of trans‐AT PKSs and provide diverse modular tools for engineered biosynthesis from a close relative of E. coli.

Here, we report the synthesis of all major 2-alkyl-4(1H)-quinolone N-oxide classes of Pseudomonas and Burkholderia, quantification of their native production levels and their antibiotic activities against competing Staphylococcus aureus.

Genome mining and bioactivity studies suggested the sponge‐derived bacterium Aquimarina sp. Aq135 as a producer of new antibiotics. Activity‐guided isolation identified antibacterial peptides, named aquimarins, featuring a new scaffold with an unusual C‐terminal amino group and chlorine moieties. Responsible for the halogenation is the FeII/α‐ketoglutarate‐dependent chlorinase AqmA that halogenates up to two isoleucine residues in a carrier protein‐dependent fashion. Total syntheses of two natural aquimarins and eight non‐natural variants were developed. Structure–activity relationship (SAR) studies with these compounds showed that the synthetically more laborious chlorinations are not required for antibacterial activity but enhance cytotoxicity. In contrast, variants lacking the C‐terminal amine were virtually inactive, suggesting diamines similar to the terminal aquimarin residue as candidate building blocks for new peptidomimetic antibiotics.