Crystal structure of colicin E3 immunity protein: an inhibitor of a ribosome-inactivating RNase

Background: Colicins are antibiotic-like proteins of Escherichia. coli that kill related strains. Colicin E3 acts as an RNase that specifically cleaves 1 6S rRNA, thereby inactivating the ribosomes in the infected cell. The producing organism is protected against colicin E3 by a specific inhibitor, the immunity protein lm3, which forms a tight 1:1 complex with colicin E3 and renders it inactive. Crystallographic studies on colicin E3 and lm3 have been undertaken to unravel the structural basis for the ribonucleolytic activity and its inhibition. Results: The crystal structure of lm3 has been determined to a resolution of 1.8 Angstrom. The structure consists of a four-stranded antiparallel beta sheet flanked by three alpha helices on one side of the sheet. Thr7, Phe9, Phe16 and Phe74 form a hydrophobic cluster on the surface of the protein in the vicinity of Cys47. This cluster is part of a putative binding pocket which also includes nine polar residues. Conclusions: The putative binding pocket of lm3 is the probable site of interaction with colicin E3. The six acidic residues in the pocket may interact with some of the numerous basic residues of colicin E3. The involvement of hydrophobic moieties in the binding is consistent with the observation that the tight complex can only be dissociated by denaturation. The structure of lm3 resembles those of certain nucleic acid binding proteins, in particular domain II of topoisomerase I and RNA-binding proteins that contain the ribonucleoprotein (RNP) sequence motif. This observation suggests that lm3 has a nucleic acid binding function in addition to binding colicin E3.