Protective Bacterial Toxin TisB Produces Well-Defined Anion-Selective Pores in Planar Lipid Bilayers

Abstract

Recently identified small bacterial peptide TisB is a component of a toxin/antitoxin system. TisB toxin induces formation of drug-tolerant persister cells in response to DNA damage. We have found that TisB forms well-defined ion-conductive pores in planar lipid bilayers. Using high-resolution conductance recordings with membranes of varying lipid compositions, we show that bath solution concentrations of TisB higher than 10 µM induce multilevel conductive states, which resemble pore formation by the well-known channel-former antibiotic alamethicin. In 1 M KCl TisB-induced pores usually first appear as stable conductive ohmic states (0.5, 1.5, and 2.6 nS), and, as time progresses, tend to produce various higher conductive states. The transition to these states is also favored by application of higher positive or negative transmembrane voltages. Both low and high conductive states possess close anion selectivity (∼80 % anion current, measured in 1M/0.1M KCl salt gradient). Probing TisB pores in their lowest conductive states with differently-sized polyethylene glycols (PEGs) shows only minute polymer partitioning even for the smallest PEGs of 200 and 300 molecular weight. This finding implies that the lowest conductive states are characterized by relatively small diameter of the aqueous pores. TisB apparently creates a dormant state in persister cells by decreasing the protein motive force across their membranes and reducing ATP production, which leads to antibiotic tolerance.