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. 2021 Jul 8;10(7):830. doi: 10.3390/antibiotics10070830

Figure 2.

Figure 2

Characterization of AcrB mutants defective in substrate binding. (a) The point mutation did not affect expression level. Anti-AcrB Western blot analysis of basal expression of all three mutants and the wild type AcrB from plasmid transformed into BW25113. Sample prepared from plasmid-free BW25113 (\) was also prepared and loaded to serve as a control to highlight the difference in expression levels. (b) Anti-AcrB and Anti-AcrA Western blot analyses revealing the formation of disulfide bonded AcrA-AcrB complexes, which was reduced after incubation with BME. AcrA-P57C/AcrB-N191C, F610A is in lane 1 and 3, and AcrA-T217C/AcrB-S258C, F610A in lane 2 and 4. Molecular weight markers are labeled as “M” and the molecular weight of bands (kD) were indicated on the right. The expected bands for AcrA, AcrB, and disulfide bond linked AcrA-AcrB are marked on the left of the gels as A, B, and AB, respectively.