Enzymatic C-Acylation Enables Diversification of Tetramate Antibiotic Scaffolds

Tetramate (pyrrolidine-2,4-dione) scaffolds are widely distributed among bacterial natural products, including metabolites produced by human-associated microorganisms, where they contribute to microbial competition and ecological fitness. Enzymatic tailoring reactions play important roles in diversifying these bioactive scaffolds; however, enzymatic C-acylation has thus far been largely restricted to Friedel–Crafts-type reactions on aromatic substrates. Here, we report the identification and characterization of a family of bacterial acyltransferases from distinct bacterial species, including Streptococcus macacae (SmaATase), Lactococcus hircilactis (LhATase), Priestia megaterium (PmATase), and Bacillus pseudomycoides (BpATase). These enzymes exhibit broad tolerance toward tetramate acceptors and phenyl acyl ester donors, enabling enzymatic diversification beyond classical aromatic frameworks. Selected acylated products retained or modestly improved antibacterial activity against methicillin-resistant Staphylococcus aureus, highlighting the potential of this enzymatic platform for expanding the chemical space of tetramate metabolites. Together, this work establishes an enzymatic strategy for diversification of tetramate natural product scaffolds and expands the scope of enzymatic C–C bond formation in microbial natural product chemistry.