(228w) Structural Basis of Macrocyclization in a Fungal Nonribosomal Peptide through a Condensation-like Termination Domain

Nonribosomal peptide synthetases (NRPSs) in fungi biosynthesize some of the most important pharmaceutical compounds, including penicillin, cyclosporine and echinocandin. The key difference between fungal and bacterial NRPSs is the use of a condensation like (C_T) domain instead of the canonical thioesterase (TE) domain in catalyzing the macrocyclization reaction. In contrast to the TE domain, the C_T domain does not form covalent intermediate during catalysis and requires specific protein-protein interactions with the upstream T domain. To understand the fungal strategy of forging the macrocyclic peptide linkage, we determined the crystal structures of the C_T domain and the holo T-C_T complex of P. aethiopicum TqaA, a trimodular NRPS involved in fumiquinazoline F biosynthesis. Our work revealed unique structural elements in C_T that differentiates from the canonical C domains, including structural elements that seal off the acceptor site and prevent solvent entry or further peptidyl chain elongation. Solution small angle scattering (SAXS) analysis revealed phosphopanthetheine prosthetic attachment in the T domain modulates the dynamic interactions between the T and C_T domains. The structure of the TqaA C_T domain represents the first C or C-like domain determined for a fungal NRPS, and provides a molecule blueprint for engineering the sizes and structures of this large family of natural products.