aurovertin-b and Myocardial-Ischemia

The mitochondrial F1F0 ATP synthase is a critical enzyme that works by coupling the proton motive force generated by the electron transport chain via proton transfer through the F0 or proton-pore forming domain of this enzyme to release ATP from the catalytic F1 domain. This enzyme is regulated by calcium, ADP, and inorganic phosphate as well as increased transcription through several pathways. This enzyme is also an ATP hydrolase under ischemic conditions. This "inefficient" hydrolysis of ATP consumes 90% of ATP consumed during ischemia as shown with non-selective ATPase inhibitors oligomycin and Aurovertin B. A benzopyran analog, BMS-199264, selectively inhibits F1F0 ATP hydrolase activity with no effect on ATP synthase activity. BMS-199264 had no effect on ATP before ischemia, but reduced the decline in ATP during ischemia. Selective hydrolase inhibition seen with the small molecule BMS-199264 suggests a conformational change in the F1F0 ATPase enzyme when switching from synthase to hydrolase activity.

Topics: Adenosine Triphosphate; Animals; Aurovertins; Enzyme Inhibitors; Forecasting; Humans; Hydrolysis; Imidazoles; Mitochondria; Mitochondrial Proton-Translocating ATPases; Models, Biological; Myocardial Ischemia; Oligomycins; Signal Transduction

The mitochondrial F1F0 ATP synthase is responsible for the majority of ATP production in mammals and does this through a rotary catalytic mechanism. Studies show that the F1F0 ATP synthase can switch to an ATP hydrolase, and this occurs under conditions seen during myocardial ischemia. This ATP hydrolysis causes wasting of ATP that does not produce work. The degree of ATP inefficiently hydrolyzed during ischemia may be as high as 50-90% of the total. A naturally occurring, reversible inhibitor (IF-1) of the hydrolase activity is in the mitochondria, and it has a pH optimum of 6.8. Based on studies with the nonselective (inhibit both synthase and hydrolase activity) inhibitors aurovertin B and oligomycin B reduce the rate of ATP depletion during ischemia, showing that IF-1 does not completely block hydrolase activity. Oligomycin and aurovertin cannot be used for treating myocardial ischemia as they will reduce ATP production in healthy tissue. We generated a focused structure-activity relationship, and several compounds were identified that selectively inhibited the F1F0 ATP hydrolase activity while having no effect on synthase function. One compound, BMS-199264 had no effect on F1F0 ATP synthase function in submitochondrial particles while inhibiting hydrolase function, unlike oligomycin that inhibits both. BMS-199264 selectively inhibited ATP decline during ischemia while not affecting ATP production in normoxic and reperfused hearts. BMS-191264 also reduced cardiac necrosis and enhanced the recovery of contractile function following reperfusion. These data also suggest that the reversal of the synthase and hydrolase activities is not merely a chemical reaction run in reverse.

Topics: Animals; Aurovertins; Enzyme Inhibitors; Imidazoles; Mitochondria; Mitochondrial Proton-Translocating ATPases; Myocardial Ischemia; Oligomycins; Prokaryotic Initiation Factor-1; Proton-Translocating ATPases; Structure-Activity Relationship

Mitochondrial F(1)F(0)-ATPase normally synthesizes ATP in the heart, but under ischemic conditions this enzyme paradoxically causes ATP hydrolysis. Nonselective inhibitors of this enzyme (aurovertin, oligomycin) inhibit ATP synthesis in normal tissue but also inhibit ATP hydrolysis in ischemic myocardium. We characterized the profile of aurovertin and oligomycin in ischemic and nonischemic rat myocardium and compared this with the profile of BMS-199264, which only inhibits F(1)F(0)-ATP hydrolase activity. In isolated rat hearts, aurovertin (1-10 microM) and oligomycin (10 microM), at concentrations inhibiting ATPase activity, reduced ATP concentration and contractile function in the nonischemic heart but significantly reduced the rate of ATP depletion during ischemia. They also inhibited recovery of reperfusion ATP and contractile function, consistent with nonselective F(1)F(0)-ATPase inhibitory activity, which suggests that upon reperfusion, the hydrolase activity switches back to ATP synthesis. BMS-199264 inhibits F(1)F(0) hydrolase activity in submitochondrial particles with no effect on ATP synthase activity. BMS-199264 (1-10 microM) conserved ATP in rat hearts during ischemia while having no effect on preischemic contractile function or ATP concentration. Reperfusion ATP levels were replenished faster and necrosis was reduced by BMS-199264. ATP hydrolase activity ex vivo was selectively inhibited by BMS-199264. Therefore, excessive ATP hydrolysis by F(1)F(0)-ATPase contributes to the decline in cardiac energy reserve during ischemia and selective inhibition of ATP hydrolase activity can protect ischemic myocardium.

Topics: Adenosine Triphosphate; Animals; Aurovertins; Cell Survival; Enzyme Inhibitors; Hydrolysis; Imidazoles; Male; Mitochondria; Myocardial Ischemia; Myocardium; Oligomycins; Proton-Translocating ATPases; Rats; Rats, Sprague-Dawley; Uncoupling Agents