Lauquin G J, Devaux P F, Bienvenüe A, Villiers C, Vignais P V
Biochemistry. 1977 Mar 22;16(6):1202-8. doi: 10.1021/bi00625a027.
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.
已合成了多种苍术苷的自旋标记酰基衍生物，即(m,n)酰基 - ATR（通式：CH3 - (CH2)mCX(CH2)nCOO - ATR，其中X为含氮氧自由基的邻唑烷环）。如自旋标记酰基 - ATR的电子自旋共振（ESR）光谱所示，当掺入脂质体中时，位于酰基链上的氮氧自由基与苍术苷部分的二萜残基相互作用。自旋标记酰基 - ATR用于探测心脏线粒体中的ADP载体。它们抑制ADP转运的效率与未标记的酰基 - ATR相同。这种抑制是混合竞争性和非竞争性抑制。抑制常数接近10(-7) M。长链酰基 - ATR（(10,3)-、(7,6)-、(7,8)-和(5,10)酰基 - ATR）以及短链(0,2)酰基 - ATR，当以低浓度添加到心脏线粒体中时，比添加到脂质体中产生更多固定化的ESR光谱。该系列的前三个分子的固定化更强。(1,14)酰基 - ATR在酰基链末端甲基附近几乎带有一个氮氧自由基，产生对应于高度流动性的光谱。加入苍术苷或其他特异性配体后，与ADP载体结合的自旋标记长链酰基 - ATR从其结合位点向线粒体膜的脂质相位移，短链(0,2)酰基 - ATR释放到水相中。当与“外翻”亚线粒体颗粒一起孵育时，自旋标记长链酰基 - ATR没有显示出与蛋白质结合的任何证据，尽管这些颗粒能够转运ADP。就线粒体膜中ADP载体的大小和不对称性以及ADP转运机制对这些结果进行了讨论。
