Bacteria use several different secretion systems to deliver toxic EndoU ribonucleases into neighboring cells. Here, we present the first structure of a prokaryotic EndoU toxin in complex with its cognate immunity protein. The contact-dependent growth inhibition toxin CdiA-CT STECO31 from Escherichia coli STEC_O31 adopts the eukaryotic EndoU fold and shares greatest structural homology with the nuclease domain of coronavirus Nsp15. The toxin contains a canonical His-His-Lys catalytic triad in the same arrangement as eukaryotic EndoU domains, but lacks the uridylate-specific ribonuclease activity that characterizes the superfamily. Comparative sequence analysis indicates that bacterial EndoU domains segregate into at least three major clades based on structural variations in the N-terminal subdomain. Representative EndoU nucleases from clades I and II degrade tRNA molecules with little specificity. In contrast, CdiA-CT STECO31 and other clade III toxins are specific anticodon nucleases that cleave tRNA Glu between nucleotides C37 and m 2 A38. These findings suggest that the EndoU fold is a versatile scaffold for the evolution of novel substrate specificities. Such functional plasticity may account for the widespread use of EndoU effectors by diverse inter-bacterial toxin delivery systems.
Bibliographical noteFunding Information:
We would like to thank Gyorgy Babnigg for help with construct design. This work was supported by National Institutes of Health grants GM102318 (C.W.G., C.S.H., D.A.L. & subcontract to A.J.), GM117373 (C.W.G., D.A.L., C.S.H.), GM094585 and GM115586 (A.J.) and the U. S. Department of Energy, Office of Biological and Environmental Research, under contract DE-AC02-06CH11357 (A.J.).
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