Diversification of β-Augmentation Interactions between CDI Toxin/Immunity Proteins

Contact-dependent growth inhibition (CDI) is a widespread mechanism of inter-bacterial competition mediated by the CdiB/CdiA family of two-partner secretion proteins. CdiA effectors carry diverse C-terminal toxin domains (CdiA-CT), which are delivered into neighboring target cells to inhibit growth. CDI+ bacteria also produce Cdil immunity proteins that bind specifically to cognate CdiA-CT toxins and protect the cell from auto-inhibition. Here, we compare the structures of homologous CdiA-CT/Cdil complexes from Escherichia coli EC869 and Yersinia pseudotuberculosis YPIII to explore the evolution of CD! toxin/immunity protein interactions. Both complexes share an unusual beta-augmentation interaction, in which the toxin domain extends a beta-hairpin into the immunity protein to complete a six-stranded anti-parallel sheet. However, the specific contacts differ substantially between the two complexes. The EC869 beta-hairpin interacts mainly through direct H-bond and ion-pair interactions, whereas the YPIII beta-hairpin pocket contains more hydrophobic contacts and a network of bridging water molecules. In accord with these differences, we find that each Cdil protein only protects target bacteria from its cognate CdiA-CT toxin. The compact beta-hairpin binding pocket within the immunity protein represents a tractable system for the rationale design of small molecules to block CdiA-CT/Cdil complex formation. We synthesized a macrocyclic peptide mimic of the beta-hairpin from EC869 toxin and solved its structure in complex with cognate immunity protein. These latter studies suggest that small molecules could potentially be used to disrupt CD! toxin/immunity complexes. (C) 2015 Elsevier Ltd. All rights reserved.