Brenza J M, Neagle C E, Sokolove P M
Biochem Pharmacol. 1985 Dec 15;34(24):4291-8. doi: 10.1016/0006-2952(85)90287-4.
The interaction of cardiolipin-containing, unilamellar liposomes with Ca2+ was assessed by flow dialysis in the presence of 2-100 microM 45Ca2+, using vesicles formed from phosphatidylcholine (PC) and from PC and cardiolipin in mole ratios from 16:1 to 1:1. Control (PC only) vesicles bound no detectable Ca2+. In contrast, Ca2+ binding to cardiolipin-containing vesicles was substantial and dependent on vesicle concentration. Scatchard plots for the binding were concave upward. Resolution of the data, assuming the presence of two independent classes of binding sites, indicated a high-affinity site with apparent KD = 5.57 +/- 0.48 microM (S.D.) and a second site with KD in the millimolar range. Interaction of cardiolipin-containing liposomes with Ca2+ was insensitive to monovalent cations (Na+, K+, Rb+), but was inhibited by ruthenium red much greater than La3+ greater than Mn2+ greater than Mg2+. Progressive increases in the PC: cardiolipin ratio markedly increased the apparent KD for Ca2+ at the high-affinity site. Stoichiometry of Ca2+ binding at the site passed through a maximum at a PC: cardiolipin ratio of 4:1. The potent antineoplastic agent adriamycin also inhibited the interaction of Ca2+ with cardiolipin-containing liposomes in a dose-dependent manner; effects were detected at 10 microM antibiotic. Unlike PC, adriamycin altered the stoichiometry of the high-affinity interaction but not the apparent KD. Adriamycin effects increased with pH in the range of the pKA of its amino group. These results suggest that inhibition by adriamycin may result from a mechanism other than simple competition for the charged head group of cardiolipin.
通过流动透析法，在2 - 100微摩尔45Ca2+存在的情况下，评估了含心磷脂的单层脂质体与Ca2+的相互作用。所用脂质体由磷脂酰胆碱（PC）以及PC与心磷脂以16:1至1:1的摩尔比形成。对照（仅PC）脂质体未检测到结合Ca2+。相比之下，含心磷脂的脂质体对Ca2+有大量结合，且依赖于脂质体浓度。结合的Scatchard图向上凹。假设存在两类独立的结合位点对数据进行解析，结果表明存在一个高亲和力位点，其表观解离常数KD = 5.57 ± 0.48微摩尔（标准差），另一个位点的KD在毫摩尔范围内。含心磷脂的脂质体与Ca2+的相互作用对单价阳离子（Na+、K+、Rb+）不敏感，但钌红的抑制作用远大于La3+大于Mn2+大于Mg2+。PC与心磷脂比例的逐渐增加显著提高了高亲和力位点处Ca2+的表观KD。该位点Ca2+结合的化学计量在PC与心磷脂比例为4:1时达到最大值。强效抗肿瘤药物阿霉素也以剂量依赖方式抑制Ca2+与含心磷脂脂质体的相互作用；在10微摩尔抗生素浓度下即可检测到作用。与PC不同，阿霉素改变了高亲和力相互作用的化学计量，但未改变表观KD。在其氨基的pKA范围内，阿霉素的作用随pH升高而增强。这些结果表明，阿霉素的抑制作用可能源于一种不同于简单竞争心磷脂带电头部基团的机制。
