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Deletion of a unique loop in the mycobacterial F‐ ATP synthase γ subunit sheds light on its inhibitory role in ATP hydrolysis‐driven H + pumping
Author(s) -
Hotra Adam,
Suter Manuel,
Biuković Goran,
Ragunathan Priya,
Kundu Subhashri,
Dick Thomas,
Grüber Gerhard
Publication year - 2016
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/febs.13715
Subject(s) - atp synthase , atp hydrolysis , mycobacterium smegmatis , atp synthase gamma subunit , biochemistry , protein subunit , atpase , enzyme , mutant , chemistry , biology , mycobacterium tuberculosis , gene , medicine , tuberculosis , pathology
The F 1 F O ‐ ATP synthase is one of the enzymes that is essential to meet the energy requirement of both the proliferating aerobic and hypoxic dormant stages of the life cycle of mycobacteria. Most F‐ ATP synthases consume ATP in the α 3 :β 3 headpiece to drive the γ subunit, which couples ATP cleavage with proton pumping in the c ring of F O via the bottom of the γ subunit. ATP ase‐driven H + pumping is latent in mycobacteria. The presence of a unique 14 amino acid residue loop of the mycobacterial γ subunit has been described and aligned in close vicinity to the c ‐ring loop Priya R et al . (2013) J Bioenerg Biomembr 45, 121‐129 Here, we used inverted membrane vesicles ( IMV s) of fast‐growing Mycobacterium smegmatis and a variety of covalent and non‐covalent inhibitors to characterize the ATP hydrolysis activity of the F‐ ATP synthase inside IMV s. These vesicles formed a platform to investigate the function of the unique mycobaterial γ loop by deleting the respective loop‐encoding sequence (γ 166–179 ) in the genome of M. smegmatis . ATP hydrolysis‐driven H + pumping was observed in IMV s containing the Δγ 166–179 mutant protein but not for IMV s containing the wild‐type F‐ ATP synthase. In addition, when compared to the wild‐type enzyme, IMV s containing the Δγ 166–179 mutant protein showed increased ATP cleavage and lower levels of ATP synthesis, demonstrating that the loop affects ATP ase activity, ATP ase‐driven H + pumping and ATP synthesis. These results further indicate that the loop may affect coupling of ATP hydrolysis and synthesis in a different mode.