Experimental measurements of conductive properties of MscS in native spheroplast membranes in a wide range of membrane potentials. (A) A typical response of channel population to a ramp of pressure at +40 mV pipette voltage. Single-channel steps are seen in the magnified trace. (B) Single-channel I-V curves obtained in symmetrical 0.2 and 1.5 M KCl in excised inside-out patches. The currents were normalized to the specific bulk conductivites of the recording buffer. Current–voltage relationships measured in an extended range of voltages using the “pulse” regimen. (C) Ensemble current measurements. A response of a patch to a train of 50-ms pulses. The pipette pressure was gradually ramped up to a saturating level, and then a train of voltage pulses was applied. The magnified top part of one current response was fit with an exponent, whose extrapolation shows the magnitude of the current at the onset of the pulse. (D) I-V plots for WT MscS obtained by the single-channel (filled symbols) and ensemble (open symbols) methods in 0.2 and 1.5 M KCl. (E) I-V plots for WT and K169Q MscS measured with the single-channel method. The inset shows the entire curves, whereas the main panel presents the magnified view of the lower left quadrant where the WT and K169Q curves deviate the most. The small error bars represent standard deviation based on three independent experiments.