Membrane Changes in Escherichia Coli Induced by Colicin Ia and Agents Known to Disrupt Energy Transduction

The addition of colicin la to a suspension of intact Escherichia coli in the presence of the hydrophobic fluorescent probe N-phenyl-1-naphthylamine causes dramatic changes in the fluorescence of the probe. The fluorescence intensity increases several fold, the emission spectrum shifts to the blue, the fluorescence lifetime approximately doubles, and the polarization increases. These changes do not appear to result from an increase in membrane microviscosity, as has been previously postulated to be the case for the N-phenyl-1-naphthylamine fluorescence changes seen with colicin E1-treated cells (Helgerson, S. L, Cramer, W. A., Harris, J. M., and Lytle, F. E. (1974), Biochemistry, 13, 3057); rather, they result from an increased binding of the dye to the cell envelope. A variety of agents have been used todemonstrate that a very similar fluorescence response results whenever the cells are “deenergized.” These agents include electron transport inhibitors (malonate, amytal, cyanide) as well as the uncouplers CCCP and azide. In addition, depleting the cells of either endogenous substrates or oxygen results in the same fluorescence response. In these cases, the fluorescence response is reversed upon addition of an oxidizable substrate or oxygen. It is clear that there are significant changes in the Escherichia coli envelope as energy transduction processes are disrupted and restored. The changes reported by the fluorescent probe may prove useful in deciphering structure-function relationships in the Escherichia coli envelope.