Communication Breakdown: Dissecting the COM Interfaces between the Subunits of Nonribosomal Peptide Synthetases

Nonribosomal peptides are a structurally diverse and bioactive class of natural products constructed by multidomain enzymatic assembly lines known as nonribosomal peptide synthetases (NRPSs). While the core catalytic domains even entire protein subunits NRPSs have been elucidated, little biophysical work has reported on docking that promote interactions—and thus transfer biosynthetic intermediates—between subunits. In present study, we closely examine COM mediate COMmunication between donor epimerization (E) acceptor condensation (C) found at termini NRPS Through combination X-ray crystallography, circular dichroism spectroscopy, solution- solid-state NMR molecular dynamics (MD) simulations, provide direct evidence for an intrinsically disordered region folds into dynamic helical motif upon binding to suitable acceptor. Furthermore, our titration carbene footprinting experiments illuminate residues involved interaction interface, MD simulations demonstrate folding consistent with experimental data. Although results lend credence previously proposed helix-hand mode interaction, they also underscore importance viewing interfaces ensembles rather than single rigid structures suggest engineering should account interactions which transiently guide in addition those stabilize final complex. activity assays affinity measurements, further substantiate role C domain implicate this short readily transposable noncognate crosstalk. Finally, bioinformatics analyses show widespread product pathways function beyond canonical type described above, setting high priority thorough characterization these domains. Our findings lay groundwork future attempts rationally engineer domain–domain ultimate goal generating molecules.