Targeting Mycobacterial F-ATP Synthase C-Terminal α Subunit Interaction Motif on Rotary Subunit γ

Mycobacteria regulate their energy (ATP) levels to sustain their survival even in stringent living conditions. Recent studies have shown that mycobacteria not only slow down their respiratory rate but also block ATP hydrolysis of the F-ATP synthase (alpha(3):beta(3):gamma:delta:epsilon:a:b:b':c(9)) to maintain ATP homeostasis in situations not amenable for growth. The mycobacteria-specific alpha C-terminus (alpha 533-545) has unraveled to be the major regulative of latent ATP hydrolysis. Its deletion stimulates ATPase activity while reducing ATP synthesis. In one of the six rotational states of F-ATP synthase, alpha 533-545 has been visualized to dock deep into subunit gamma, thereby blocking rotation of gamma within the engine. The functional role(s) of this C-terminus in the other rotational states are not clarified yet and are being still pursued in structural studies. Based on the interaction pattern of the docked alpha 533-545 region with subunit gamma, we attempted to study the druggability of the alpha 533-545 motif. In this direction, our computational work has led to the development of an eight-featured alpha 533-545 peptide pharmacophore, followed by database screening, molecular docking, and pose selection, resulting in eleven hit molecules. ATP synthesis inhibition assays using recombinant ATP synthase as well as mycobacterial inverted membrane vesicles show that one of the hits, AlMF1, inhibited the mycobacterial F-ATP synthase in a micromolar range. The successful targeting of the alpha 533-545-gamma interaction motif demonstrates the potential to develop inhibitors targeting the alpha site to interrupt rotary coupling with ATP synthesis.

Automated summary

Recent studies have shown that mycobacteria regulate their ATP levels by slowing down respiratory rate and blocking ATP hydrolysis of the F-ATP synthase, with the alpha 533-545 region identified as a major regulator. The alpha 533-545 motif has been found to inhibit rotary coupling with ATP synthesis by docking deep into subunit gamma of the enzyme.