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The mitochondrial permeability transition pore (mPTP), a high-conductance channel triggered by a sudden Ca 2+ concentration increase, is composed of the F 1 F O -ATPase. Since mPTP opening leads to mitochondrial dysfunction, which is a feature of many diseases, a great pharmacological challenge is to find mPTP modulators. In our study, the effects of two 1,5-disubstituted 1,2,3-triazole derivatives, five-membered heterocycles with three nitrogen atoms in the ring and capable of forming secondary interactions with proteins, were investigated. Compounds 3a and 3b were selected among a wide range of structurally related compounds because of their chemical properties and effectiveness in preliminary studies. In swine heart mitochondria, both compounds inhibit Ca 2+ -activated F 1 F O -ATPase without affecting F-ATPase activity sustained by the natural cofactor Mg 2+ . The inhibition is mutually exclusive, probably because of their shared enzyme site, and uncompetitive with respect to the ATP substrate, since they only bind to the enzyme-ATP complex. Both compounds show the same inhibition constant (K' i ), but compound 3a has a doubled inactivation rate constant compared with compound 3b. Moreover, both compounds desensitize mPTP opening without altering mitochondrial respiration. The results strengthen the link between Ca 2+ -activated F 1 F O -ATPase and mPTP and suggest that these inhibitors can be pharmacologically exploited to counteract mPTP-related diseases., (© 2020 New York Academy of Sciences.)

The properties of the mitochondrial F 1 F O -ATPase catalytic site, which can bind Mg 2+ , Mn 2+ , or Ca 2+ and hydrolyze ATP, were explored by inhibition kinetic analyses to cast light on the Ca 2+ -activated F 1 F O -ATPase connection with the permeability transition pore (PTP) that initiates cascade events leading to cell death. While the natural cofactor Mg 2+ activates the F 1 F O -ATPase in competition with Mn 2+ , Ca 2+ is a noncompetitive inhibitor in the presence of Mg 2+ . Selective F 1 inhibitors (Is-F 1 ), namely NBD-Cl, piceatannol, resveratrol, and quercetin, exerted different mechanisms (mixed and uncompetitive inhibition) on either Ca 2+ - or Mg 2+ -activated F 1 F O -ATPase, consistent with the conclusion that the catalytic mechanism changes when Mg 2+ is replaced by Ca 2+ . In a partially purified F 1 domain preparation, Ca 2+ -activated F 1 -ATPase maintained Is-F 1 sensitivity, and enzyme inhibition was accompanied by the maintenance of the mitochondrial calcium retention capacity and membrane potential. The data strengthen the structural relationship between Ca 2+ -activated F 1 F O -ATPase and the PTP, and, in turn, on consequences, such as physiopathological cellular changes., (© 2019 New York Academy of Sciences.)