Site-Directed Mutagenesis Reveals Amino Acid Residues in the Escherichia coli RND Efflux Pump AcrB That Confer Macrolide Resistance

Macrolide resistance mechanisms inEnterobacteriaceae: Focus on azithromycin

“…In a study on Salmonella enterica serovar Typhimurium with an inactivated acrB gene, Nikaido et al (1998) observed a 64-fold decrease in the MIC of erythromycin (from 512 mg/L to 8 mg/L). These observations are consistent with the aforementioned effect of the PAbN (Gomes et al, 2013b) and with the 128-fold decreases in the MIC levels to erythromycin, clarithromycin and azithromycin (from 512 mg/L to 4 mg/L for the first two and from 64 to 0.5 for azithromycin) described by Wehmeier et al (2009) when disrupting the acrB gene. As with other antimicrobial families, this kind of generic efflux pumps are not able to extrude all macrolide members from the bacterial cytoplasm, thereby demonstrating the inability of AcrAB-TolC to pump out telithromycin (Chollet et al, 2004).…”

“…However, not all alterations in either 23S rRNA or the L4 and L22 proteins result in cross-resistance to all members of the macrolide family, with some being affected by specific alterations (Ettayebi et al, 1985). Chromosomal efflux pumps have been reported to have a relevant role in macrolides resistance (Gomes et al, 2013b;Wehmeier et al, 2009). In addition, a series of less studied chromosomal encoded mechanisms, such as the presence of short peptides encoded in "mini-genes" able to affect the susceptibility levels to macrolides have also been described Tripathi et al, 1998) despite not taking part in the development of macrolide resistance in clinical Enterobacteriaceae.…”

Quinazoline derivatives are efficient chemosensitizers of antibiotic activity in Enterobacter aerogenes, Klebsiella pneumoniae and Pseudomonas aeruginosa resistant strains

“…Moreover, several recent data on the E. coli AcrB efflux mechanism indicate that some amino acid residues are involved in the transport process to a varying extent during specific steps such as binding, transport, etc. [14][15][16]. This can explain the divergence observed in the competitive capacity of the compounds with drugs of different classes (chloramphenicol, quinolones, ethidium bromide) for binding sites within the pump type (AcrB, MexB) as well as the level of expressed efflux pump activity in the tested strains.…”

“…Kinetic studies have reported variation in the affinity constant of various -lactams for AcrB in E. coli cells [13]. This flexibility of the substrate site may be directly involved in the polyselectivity of the efflux pump; a site that can be modified by site-directed mutagenesis [15,16]. The role and overlap of affinity sites may explain the discrepancy observed in the increase of intracellular concentrations of different transported molecules such as chloramphenicol, quinolones and -lactams.…”

Mechanism of recognition of compounds of diverse structures by the multidrug efflux pump AcrB ofEscherichia coli

“…Phe178Ala mutation, on the wall of the groove, also lowered the MICs of groove-binders such as novobiocin and erythromycin, but mutations of the residues on the walls of the cave (Phe136, Phe615) had little effect on MICs of the cave-binders such as NMP and ethidium, perhaps because the cave area is so wide. Phe615Ala mutation unexpectedly affected only the MICs of macrolides, and a later study (38) showed the effect of 615-617 area on macrolide specificity. Although erythromycin was predicted to bind predominantly to the groove area, parts of the large molecule may spill over to the cave area (Fig.…”