Macrolides are potent inhibitors of FO sector of the mitochondrial F1FO-ATPase. The present work explores the interaction mechanism(s) of natural macrolide antibiotics with FO in swine heart mitochondria. Oligomycin, venturicidin and bafilomycin, which share the macrolide ring and differ in some side chains, are uncompetitive inhibitors of the mitochondrial ATPase. Kinetic data from the ATPase inhibition by binary mixtures of macrolides (I1+I2) suggest that any combination of two out of three macrolides tested can yield the quaternary complex ESI1I2 through non-overlapping binding sites and produce synergistic inhibition. The high F1FO-ATPase sensitivity to macrolides is strongly decreased by the known mitochondrial poison tributyltin (TBT) at concentrations >1 μM. Accordingly, the ampiphilic TBT dose-dependently produces an exponential increase in the macrolideinsensitive ATPase activity. Since the enzyme desensitization by TBT is overcome by the reducing agent dithioerythritol (DTE), critical thiol groups are likely to be involved in TBT binding. Data suggest that the onset of covalent tin-sulphur bonds, which implies thiol oxidation due to TBT binding, would alter the interhelical packing of protomers and destabilize the interactions between the macrolide(s) and c-ring. According to the hypothesized mechanism, in the presence of macrolides, the drug-inhibited enzyme conformation which prevents proton flux through the electrical rotary motor FO cannot be established. Under such conditions DTE addition, by reducing the thiol groups oxidized by TBT, would restore the native macrolide-inhibited enzyme conformation. The whole of data suggest that the various macrolides tested share the same inhibition mechanism of the F1FO-ATPase by binding to a specific site into a common drug-binding region. These findings, other than casting light on some poorly explored mechanisms of basic bioenergetics, may be potentially helpful for drug design.
Oligomycin, venturicidin and bafilomycin bind to a specific site in a common macrolide-binding region of FO-c subunits of the ATP synthase.
NESCI, SALVATORE;VENTRELLA, VITTORIA;TROMBETTI, FABIANA;PAGLIARANI, ALESSANDRA
2013
Abstract
Macrolides are potent inhibitors of FO sector of the mitochondrial F1FO-ATPase. The present work explores the interaction mechanism(s) of natural macrolide antibiotics with FO in swine heart mitochondria. Oligomycin, venturicidin and bafilomycin, which share the macrolide ring and differ in some side chains, are uncompetitive inhibitors of the mitochondrial ATPase. Kinetic data from the ATPase inhibition by binary mixtures of macrolides (I1+I2) suggest that any combination of two out of three macrolides tested can yield the quaternary complex ESI1I2 through non-overlapping binding sites and produce synergistic inhibition. The high F1FO-ATPase sensitivity to macrolides is strongly decreased by the known mitochondrial poison tributyltin (TBT) at concentrations >1 μM. Accordingly, the ampiphilic TBT dose-dependently produces an exponential increase in the macrolideinsensitive ATPase activity. Since the enzyme desensitization by TBT is overcome by the reducing agent dithioerythritol (DTE), critical thiol groups are likely to be involved in TBT binding. Data suggest that the onset of covalent tin-sulphur bonds, which implies thiol oxidation due to TBT binding, would alter the interhelical packing of protomers and destabilize the interactions between the macrolide(s) and c-ring. According to the hypothesized mechanism, in the presence of macrolides, the drug-inhibited enzyme conformation which prevents proton flux through the electrical rotary motor FO cannot be established. Under such conditions DTE addition, by reducing the thiol groups oxidized by TBT, would restore the native macrolide-inhibited enzyme conformation. The whole of data suggest that the various macrolides tested share the same inhibition mechanism of the F1FO-ATPase by binding to a specific site into a common drug-binding region. These findings, other than casting light on some poorly explored mechanisms of basic bioenergetics, may be potentially helpful for drug design.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
