Spin-labeled acyl atractyloside as a probe of the mitochondrial adenosine diphosphate carrier. Asymmetry of the carrier and direct lipid environment.
A number of spin-labeled acyl derivatives of atractyloside, (m,n)acyl-ATR (general formula: CH3- (CH2)mCX(CH2)nCOO-ATR, where X is an o-azolidine ring containing a nitroxide), have been synthesized. As shown by electron spin resonance (ESR) spectra of spin-labeled acyl-ATR, the nitroxide placed on the acyl chain interacts with the diterpene residue of the atractyloside moiety when incorporated in liposomes. Spin-labeled acyl-ATRs were used to probe the ADP carrier in heart mitochondria. They inhibit ADP transport with the same efficiency as unlabeled acyl-ATRs. The inhibition is a mixed competitive and noncompetitive inhibition. The inhibitor constant is close to 10(-7) M. The long chain acyl-ATRs (10,3)- (7,6)-, (7,8)-, and (5,10)acyl-ATRs) and also the short chain (0,2)acyl-ATR, when added at low concentrations to heart mitochondria, give rise to more immobilized ESR spectra than when added to liposomes. Immobilization is stronger for the first three molecules of the series. The (1,14)acyl-ATR, which possesses a nitroxide almost at the end of the acyl chain near the terminal methyl, gives rise to a spectrum corresponding to a high degree of fluidity. Upon addition of atractyloside or of other specific ligands, spin-labeled long-chain acyl-ATRs bound to the ADP carrier are displaced from their binding site toward the lipid phase of the mitochondrial membrane and the short chain (0,2)acyl-ATR is released into the aqueous phase. Spin-labeled long-chain acyl-ATRs do not show any evidence of binding to a protein when incubated with "inside out" submitochondrial particles, in spite of the fact that these particles are able to transport ADP. These results are discussed with respect to the size and the asymmetry of the ADP carrier in the mitochondrial membrane and the mechanism of ADP transport.
D000244:Adenosine Diphosphate / Q000378:metabolism* / Q000494:pharmacology; D000818:Animals; D001278:Atractyloside / Q000378:metabolism; D001665:Binding Sites; D001692:Biological Transport; D002417:Cattle; D004578:Electron Spin Resonance Spectroscopy; D006027:Glycosides / Q000378:metabolism; D007700:Kinetics; D008563:Membrane Lipids / Q000378:metabolism; D008566:Membranes / Q000378:metabolism / Q000648:ultrastructure; D008931:Mitochondria, Muscle / Q000378:metabolism* / Q000648:ultrastructure; D008954:Models, Biological; D008958:Models, Molecular; D008968:Molecular Conformation; D009206:Myocardium; D051381:Rats; D013113:Spin Labels
D006027:Glycosides; D008563:Membrane Lipids; D013113:Spin Labels; D001278:Atractyloside; D000244:Adenosine Diphosphate