Interactions involving the cyanine dye, diS-C3-(5), cytochrome c and liposomes and their implications for estimations of delta psi in cytochrome c oxidase-reconstituted proteoliposomes.
The interference of cytochrome c with absorption and fluorescence changes of the cyanine dye, diS-C3-(5), was investigated in the presence of liposomes and cytochrome c-oxidase reconstituted proteoliposomes. The apparent cytochrome c-dependent quenching of diS-C3-(5) fluorescence, and the associated absorbance losses in the presence of liposomes and proteoliposomes in low ionic strength media, are due to destruction of the dye caused by cytochrome c-mediated lipid peroxidation. The rate of this reaction was further enhanced in the presence of hydrogen peroxide. Even in the absence of liposomes or proteoliposomes, a cytochrome c-induced breakdown of dye was observed in the presence of hydrogen peroxide. The cytochrome c mediated absorbance and fluorescence losses of diS-C3-(5) in liposomal or proteoliposomal systems are prevented by Ca2+ and La3+ ions, by ascorbate, by high ionic strength and by the antioxidant BHT. Under these conditions, the process of lipid peroxidation mediated by cytochrome c is inhibited either directly (e.g. by BHT) or indirectly, by preventing the binding of cytochrome c to lipid vesicles. The impact of these findings upon the experimental estimation of membrane potential in aa3-reconstituted proteoliposomes is discussed.
Singh
A P
AP
Chanady
G A
GA
Nicholls
P
P
D000818:Animals; D001205:Ascorbic Acid / Q000494:pharmacology; D052160:Benzothiazoles; D002084:Butylated Hydroxytoluene / Q000494:pharmacology; D002232:Carbocyanines; D002413:Cations, Divalent; D003574:Cytochrome c Group / Q000378:metabolism*; D003576:Electron Transport Complex IV / Q000378:metabolism*; D005456:Fluorescent Dyes; D006736:Horses; D007700:Kinetics; D008081:Liposomes; D009206:Myocardium / Q000378:metabolism; D009994:Osmolar Concentration; D011510:Proteolipids / Q000378:metabolism*; D011804:Quinolines
D052160:Benzothiazoles; D002232:Carbocyanines; D002413:Cations, Divalent; D003574:Cytochrome c Group; D005456:Fluorescent Dyes; D008081:Liposomes; D011510:Proteolipids; D011804:Quinolines; C020485:proteoliposomes; D002084:Butylated Hydroxytoluene; C012944:3,3'-dipropyl-2,2'-thiadicarbocyanine; D003576:Electron Transport Complex IV; D001205:Ascorbic Acid