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Antimicrobial Agents and Chemotherapy, November 2000, p. 2969-2978, Vol. 44, No. 11
0066-4804/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Stages of Polymyxin B Interaction with the Escherichia coli Cell Envelope

Rimantas Daugelaviius,^1,2,* Elena Bakien,¹ and Dennis H. Bamford²

Department of Biochemistry and Biophysics, Vilnius University, iurlionio 21, LT-2009 Vilnius, Lithuania,¹ and Institute of Biotechnology and Department of Biosciences, FIN-00014 University of Helsinki, Helsinki, Finland²

Received 2 May 2000/Returned for modification 3 June 2000/Accepted 31 July 2000

The effects of polymyxin B (PMB) on the Escherichia coli outer (OM) and cytoplasmic membrane (CM) permeabilities were studied by monitoring the fluxes of tetraphenylphosphonium, phenyldicarbaundecaborane, and K⁺ and H⁺ ions. At concentrations between 2 and 20 µg/ml, PMB increased the OM permeability to lipophilic compounds and induced a leakage of K⁺ from the cytosol and an accumulation of lipophilic anions in the cellular membranes but did not cause the depolarization of the CM. At higher concentrations, PMB depolarized the CM, forming ion-permeable pores in the cell envelope. The permeability characteristics of PMB-induced pores mimic those of bacteriophage- and/or bacteriocin-induced channels. However, the bactericidal effect of PMB took place at concentrations below 20 µg/ml, indicating that this effect is not caused by pore formation. Under conditions of increased ionic strength, PMB made the OM permeable to lipophilic compounds and decreased the K⁺ gradient but was not able to depolarize the cells. The OM-permeabilizing effect of PMB can be diminished by increasing the concentration of Mg²⁺. The major new findings of this work are as follows: (i) the OM-permeabilizing action of PMB was dissected from its depolarizing effect on the CM, (ii) the PMB-induced ion-permeable pores in bacterial envelope were registered, and (iii) the pore formation and depolarization of the CM are not obligatory for the bactericidal action of PMB and dissipation of the K⁺ gradient on the CM.

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^* Corresponding author. Mailing address: Department of Biosciences, Biocentre 2, P.O. Box 56 (Viikinkaari 5), FIN-00014 University of Helsinki, Helsinki, Finland. Phone: 358 9 19159097. Fax: 358 9 19159098. E-mail: daugelav{at}cc.helsinki.fi.

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Antimicrobial Agents and Chemotherapy, November 2000, p. 2969-2978, Vol. 44, No. 11
0066-4804/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

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