Figure 5.

Co-purification of genomic AcrB with AcrA-his. (a) Anti-AcrB Western blot analyses of samples prepared from BW25113 or BW25113ΔacrA with or without plasmid-encoded AcrA-his. (b) Anti-AcrB Western blot analyses of samples collected after 0, 5, or 17 h of incubation following the induction of AcrA-his production in BW25113 or BW25113ΔacrA strains. Dithiobis-(succinimidyl proprionate) (DSP) crosslinking was performed to stabilize the AcrAB complex before protein purification. Reduction using dithiothreitol (DTT) breaks the disulfide bond in the linker of DSP and the complex into AcrA and AcrB subunits. Molecular weight markers are labeled as “M”, and the molecular weight of bands (kD) were indicated on the right. The expected bands for AcrB and DSP linked AcrA-AcrB are marked on the left of the gels as B and AB, respectively.