Interaction of the colicin K bactericidal toxin with components of its import machinery in the periplasm of Escherichia coli.
Colicins are bacterial antibiotic toxins produced by Escherichia coli cells and are active against E. coli and closely related strains. To penetrate the target cell, colicins bind to an outer membrane receptor at the cell surface and then translocate their N-terminal domain through the outer membrane and the periplasm. Once fully translocated, the N-terminal domain triggers entry of the catalytic C-terminal domain by an unknown process. Colicin K uses the Tsx nucleoside-specific receptor for binding at the cell surface, the OmpA protein for translocation through the outer membrane, and the TolABQR proteins for the transit through the periplasm. Here, we initiated studies to understand how the colicin K N-terminal domain (KT) interacts with the components of its transit machine in the periplasm. We first produced KT fused to a signal sequence for periplasm targeting. Upon production of KT in wild-type strains, cells became partly resistant to Tol-dependent colicins and sensitive to detergent, released periplasmic proteins, and outer membrane vesicles, suggesting that KT interacts with and titrates components of its import machine. Using a combination of in vivo coimmunoprecipitations and in vitro pulldown experiments, we demonstrated that KT interacts with the TolA, TolB, and TolR proteins. For the first time, we also identified an interaction between the TolQ protein and a colicin translocation domain.
Barnéoud-Arnoulet
Aurélie
A
Laboratoire d'Ingénierie des Systèmes Macromoléculaires, Institut de Microbiologie de la Méditerranée, Aix-Marseille Université, CNRS-UPR 9027, Marseille, France.
Gavioli
Marthe
M
Lloubès
Roland
R
Cascales
Eric
E
D003087:Colicins / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000187:drug effects*; D029968:Escherichia coli Proteins / Q000378:metabolism; D047468:Immunoprecipitation; D008565:Membrane Proteins / Q000378:metabolism; D033903:Periplasmic Proteins / Q000378:metabolism*; D011485:Protein Binding; D025941:Protein Interaction Mapping; D021381:Protein Transport
D003087:Colicins; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D033903:Periplasmic Proteins; C062093:tolA protein, E coli; C062094:tolB protein, E coli; C060792:tolQ protein, E coli; C060794:tolR protein, E coli