Structural and functional analysis of the promiscuous AcrB and AdeB efflux pumps suggests different drug binding mechanisms

Abstract

Upon antibiotic stress Gram-negative pathogens deploy resistance-nodulation-cell division-type tripartite efflux pumps. These include a H+/drug antiporter module that recognizes structurally diverse substances, including antibiotics. Here, we show the 3.5 Å structure of subunit AdeB from the Acinetobacter baumannii AdeABC efflux pump solved by single-particle cryo-electron microscopy. The AdeB trimer adopts mainly a resting state with all protomers in a conformation devoid of transport channels or antibiotic binding sites. However, 10% of the protomers adopt a state where three transport channels lead to the closed substrate (deep) binding pocket. A comparison between drug binding of AdeB and Escherichia coli AcrB is made via activity analysis of 20 AdeB variants, selected on basis of side chain interactions with antibiotics observed in the AcrB periplasmic domain X-ray co-structures with fusidic acid (2.3 Å), doxycycline (2.1 Å) and levofloxacin (2.7 Å). AdeABC, compared to AcrAB-TolC, confers higher resistance to E. coli towards polyaromatic compounds and lower resistance towards antibiotic compounds.

Comparative genomics analysis and virulence-related factors in novel Aliarcobacter faecis and Aliarcobacter lanthieri species identified as potential opportunistic pathogens

“…The identified pumps belong to the Resistance Nodulation cell Division (RND) protein superfamily, one of the most studied antiporters found in bacteria [ 57 ]. The identified genes were highly similar to acr B , encoding a multidrug efflux pump [ 58 ]. Acr B is a well-described antiporter involved in resistance to lipophilic β-lactam antibiotics, such as carbapenems and cephalosporins, fluoroquinolones, tetracyclines (including tigecycline), chloramphenicol, macrolides, trimethoprim, ethidium, rifampicin, and novobiocin [ 58 ].…”

“…The identified genes were highly similar to acr B , encoding a multidrug efflux pump [ 58 ]. Acr B is a well-described antiporter involved in resistance to lipophilic β-lactam antibiotics, such as carbapenems and cephalosporins, fluoroquinolones, tetracyclines (including tigecycline), chloramphenicol, macrolides, trimethoprim, ethidium, rifampicin, and novobiocin [ 58 ]. It is of particular interest as previous studies showed that A. butzleri strains exhibited resistance to a variety of antibiotics, where the majority of them belong to β-lactams and some to quinolones and coumarins [ 15 ].…”

“…As indicated above, RND transporters like Acr B may determine resistance to quinolones and coumarins [ 58 , 59 ]. This class of antibiotics targets bacterial DNA gyrase, type II topoisomerase, which plays an essential role in DNA replication [ 60 ].…”

Recognition of quinolone antibiotics by the multidrug efflux transporter MexB ofPseudomonas aeruginosa

“…Looking at all cluster representatives, we identified three main binding modes. The first binding mode (BM1) is located in the groove, where the central aromatic core is 3 Heat map of contacts (%) between all poses of LFX with the DP T of AcrB -Contacts within 3.5 Å found in the experimental structure (PDB Id: 7B8T 32 ) are reported in red boldface involved in a p-p stacking interaction with F178, F615 and F628, the carboxyl group faces K151 and the 7-piperazyl ring points towards the HT (Fig. 6C).…”

Proximal Binding Pocket Arg717 Substitutions in Escherichia coli AcrB Cause Clinically Relevant Divergencies in Resistance Profiles

“…Gly288 is a distal binding pocket residue ( Fig. 1b ), close to the hydrophobic pit, where multiple drugs bind in the binding monomer ( 13 , 22 , 28 – 33 ). Results are shown in Table 1 .…”