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New insights into the structural and functional involvement of the gate loop in AcrB export activity

DOI: 10.1016/j.bbapap.2017.11.003 DOI Help

Authors: Abdessamad Ababou (University of East London)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Biochimica Et Biophysica Acta (bba) - Proteins And Proteomics

State: Published (Approved)
Published: November 2017

Abstract: AcrB is a major multidrug exporter in Escherichia coli and other Gram-negative bacteria. Its gate loop, located between the proximal and the distal pockets, have been reported to play important role in the export of many antibiotics. This loop location, rigidity and interactions with substrates have led recent reports to suggest that AcrB export mechanism operates in a sequential manner. First the substrate binds the proximal pocket in the access monomer, then it moves to bind the distal pocket in the binding monomer and subsequently it is extruded in the extrusion monomer. Recently, we have demonstrated that the gate loop is not required for the binding of Erythromycin but the integrity of this loop is important for an efficient export of this substrate. However, here we show that the antibiotic susceptibilities of the same AcrB gate loop mutants for Doxorubicin were unaffected, suggesting that this loop is not required for its export, and we demonstrate that this substrate may use principally the tunnel-1, located between transmembranes 8 and 9, more often than previously reported. To further explain our findings, here we address the gate loop mutations effects on AcrB solution energetics (fold, stability, molecular dynamics) and on the in vivo efflux of Erythromycin and Doxorubicin. Finally, we discuss the efflux and the discrepancy between the structural and the functional experiments for Erythromycin in these gate loop mutants.

Journal Keywords: Bacterial multidrug resistance; AcrB-substrate interactions; AcrB export mechanism; X-ray crystallography; MD simulation

Diamond Keywords: Bacteria

Subject Areas: Biology and Bio-materials

Instruments: I02-Macromolecular Crystallography , I03-Macromolecular Crystallography , I04-Macromolecular Crystallography

Other Facilities: Swiss Light Source

Added On: 13/11/2017 10:33

Discipline Tags:

Structural biology Life Sciences & Biotech

Technical Tags:

Diffraction Macromolecular Crystallography (MX)