The cytotoxic domain of colicin E9 is a channel-forming endonuclease

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Mosbahi, Khédidja ; Lemaître, Christelle ; Mobasheri, Hamid ; Morel, Bertrand ; Lea, Edward J.A. ; Kleanthous, Colin ; Keeble, Anthony H. ; James, Richard ; Moore, Geoffrey R. (2002)

Bacterial toxins commonly translocate cytotoxic enzymes into cells using dedicated channelforming subunits or domains as conduits. We demonstrate that the small cytotoxic endonuclease domain from the bacterial toxin colicin E9 (the E9 DNase) exhibits nonvoltage- gated, channel-forming activity in planar lipid bilayers and that this activity is linked to toxin translocation into cells. A disulfide bond engineered into the DNase abolished channel activity and colicin toxicity but left endonuclease activity unaffected, with NMR experiments suggesting decreased conformational flexibility as the likely reason for these alterations. Concomitant with the reduction of the disulfide bond was the restoration of conformational flexibility, DNase channel activity and colicin toxicity. Our data suggest that endonuclease domains of colicins may mediate their own translocation across the bacterial inner membrane through an intrinsic channel activity that is dependent on structural plasticity in the protein.
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    1School of Biological Sciences, University of East Anglia, Norwich, NR4 7TJ, U.K 2Present Address: Laboratoire de Spectrométrie de Masse Bioorganique, Université Louis Pasteur, UMR/ULP CNRS 7509, ECPM 25 rue Becquerel, F-67087 Cedex 2, France.

    3Present Address: Laboratory of Membrane Biophysics, Institute of Biochemistry and Biophysics, University of Tehran, PO Box 13145-1384, IR Iran.

    4School of Chemical Sciences, University of East Anglia, Norwich, NR4 7TJ, U.K 5Division of Microbiology and Infectious Diseases, University Hospital, Queen’s Medical Centre, University of Nottingham, Nottingham NG7 2UH, U.K.

    6These authors contributed equally to this work.

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