Abstract
Bacterial energy metabolism is now recognized as a critical factor for the efficacy of antibiotics. The F-type ATPase/ATP synthase (FOF1) is a central player in cellular bioenergetics of bacteria and eukaryotes, and its potential as a selective antibiotic target has been confirmed by the success of bedaquiline in combatting multidrug-resistant tuberculosis. Venturicidin macrolides were initially identified for their antifungal properties and were found to specifically inhibit FOF1 of eukaryotes and bacteria. Venturicidins alone are not effective antibacterials but recently were found to have adjuvant activity, potentiating the efficacy of aminoglycoside antibiotics against several species of resistant bacteria. Here we discovered more complex effects of venturicidins on the ATPase activity of FOF1 in bacterial membranes from Escherichia coli and Pseudomonas aeruginosa. Our major finding is that higher concentrations of venturicidin induce time– and ATP–dependent decoupling of F1-ATPase activity from the venturicidin-inhibited, proton-transporting FO complex. This dysregulated ATPase activity is likely to be a key factor in the depletion of cellular ATP induced by venturicidins in prior studies with P. aeruginosa and Staphylococcus aureus. Further studies of how this functional decoupling occurs could guide development of new antibiotics and/or adjuvants that target the F-type ATPase/ATP synthase.
Highlights
The F-type ATPase/ATP synthase is a ubiquitous rotary motor enzyme involved in cellular bioenergetics in eukaryotes and bacteria
We report multiple effects of venturicidins on FOF1 in bacterial membranes, and these provide insights that could improve the efficacy of targeting bacterial F OF1 for developing new antibiotics and/or adjuvants for existing antibiotics
Venturicidins have higher affinity for F O when the coupled F 1 is in an active state (Fig. 7, B) rather than shifted to transiently inactive but significantly populated forms, the MgADP-inhibited or εCTD-inhibited states (Fig. 7, A)
Summary
The F-type ATPase/ATP synthase is a ubiquitous rotary motor enzyme involved in cellular bioenergetics in eukaryotes and bacteria. Venturicidin A (ventA) was found to potentiate the action of aminoglycoside antibiotics against various MDR bacterial pathogens[22]; this adjuvant activity was suggested to be due to ventA’s direct inhibition of ATP synthesis by FOF1 and the subsequent increase in PMF, which should potentiate uptake of aminoglycosides. Adding ventA or ventB to membranes induces immediate inhibition of ATP hydrolysis that, at higher inhibitor concentrations, is followed by a time-dependent increase in ATPase activity We show that the latter phase of ATPase recovery results from venturicidin-induced functional decoupling of F1-ATPase activity from the proton–transporting FO. With E. coli membranes, we show that minimizing the fraction of MgADP-inhibited enzyme significantly increases the enzyme’s affinity for ventA and ventB We discuss how these findings provide new insights into the likely mechanisms of venturicidins’ adjuvant activity for some antibiotics
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