Fig. 3.

Bodipy FL maleimide labeling of AcrB residues in the vestibule and the central cavity. N274C and K708C are positive and negative controls, respectively. The residues K29C, A39C, E95C, G97C, D99C, A100C, V382C, A385C, and S462C are in the central cavity. The residues N74C, R307C, V557C, S836C, E842C, S849C, L868C, and Q872 are in the vestibule. Each lane represents the labeling of a single cysteine residue by Bodipy FL maleimide (top) and a Coomassie stain (bottom) of the same gel. The labeling intensities (numbers below each lane) have been normalized (corrected for the amount of AcrB protein, determined with Coomassie stain), based on two independent experiments. The normalized labeling intensity in the positive control, N274C, was set as 100. Those residues showing intensities of <30 were considered negative.