Kinetic properties of the mitochondrial F 1 F O -ATPase activity elicited by Ca 2+ in replacement of Mg 2+

Mitochondrial Ca2+‐activated F1FO‐ATPase hydrolyzes ATP and promotes the permeability transition pore

“…Once the reaction was stopped, vials were centrifuged for 15 min at 3500 rpm (Eppendorf Centrifuge 5202). In the supernatant, the concentration of inorganic phosphate (Pi) hydrolyzed by known amounts of mitochondrial protein, which is an indirect measure of F‐ATPase activity, was spectrophotometrically evaluated . For this aim, 1 µL from a mother solution of 3 mg/mL oligomycin in dimethylsulfoxide (DMSO) was directly added to the reaction mixture before starting the reaction.…”

“…The F 1 F O ‐ATPase of Escherichia coli hydrolyzes ATP in the presence of Ca 2+ and forms a pH gradient with nearly the same effectiveness as Mg 2+ . In most bacteria, chloroplasts, and beef heart submitochondrial particles, Ca 2+ ‐activated F 1 F O ‐ATPase has been reported to be unable to translocate H + ; however, in swine heart mitochondria, the inhibition of Ca 2+ ‐activated F 1 F O ‐ATPase by oligomycin and DCCD (both blockers of H + translocation within F O ) is of the same extent as inhibition of Mg 2+ ‐activated F 1 F O ‐ATPase . In inside‐out submitochondrial particles, torque generation by the central stalk driven by Ca‐ATP hydrolysis in F 1 is coupled to the membrane F O sector and is sensitive to the transmembrane electrochemical H + gradient .…”

The mitochondrial permeability transition pore in cell death: A promising drug binding bioarchitecture

“…These inhibitors should also have no off‐target effects outside the mitochondrial enzyme. The known F O inhibitors (oligomycin and DCCD) by blocking the H + translocation inhibit both the Ca 2+ and Mg 2+ ‐activated F 1 F O ‐ATPase . The mPTP opening detected by calcein quenching rate was reduced by oligomycin and DCCD as efficiently as by CsA .…”

“…Even if the catalytic properties of substrate binding to the enzyme are independent of the cation that acts as cofactor, the catalytic sites may adopt different conformations if Mg 2+ is replaced by Ca 2+ . Recent findings suggest that in the latter case, the F 1 F O ‐ATPase is directly involved in the mPTP formation . Molecular dynamics simulations of the ( αβ ) 3 ‐OSCP complex in the F 1 domain strongly suggest that Ca 2+ insertion in the enzyme catalytic sites when Ca 2+ replaces Mg 2+ triggers conformational changes, which are mechanically transmitted from the hydrophilic sector F 1 to the membrane portion F O in the IMM.…”

“…In addition, small molecules mimicking the spiroketal portion of oligomycin target the c subunit(s) of F O domain and inhibit the mPTP without affecting ATP synthesis . Ca 2+ binding to the F 1 domain involves a catalytic mechanism that may be cation‐dependent . The higher Ca 2+ steric hindrance with respect to Mg 2+ (Figure ) implies the onset of a more flexible coordination geometry characterized by irregular bond distances and angles in the enzyme catalytic sites of β subunit.…”