Powell G L, Knowles P F, Marsh D
Abteilung Spektroskopie, Max-Planck-Institut für biophysikalische Chemie, Göttingen, Federal Republic of Germany.
Biochemistry. 1987 Dec 15;26(25):8138-45. doi: 10.1021/bi00399a018.
The selectivity of interaction of various cardiolipin analogues with beef heart cytochrome oxidase in reconstituted complexes with dimyristoylphosphatidylcholine has been studied by electron spin resonance spectroscopy, using lipids spin-labeled in the acyl chains. No difference in selectivity is observed between cardiolipin and its monolyso derivative, and similarly no selectivity is observed between phosphatidylcholine and lysophosphatidylcholine. Removal of the cardiolipin charge by methylation of the phosphate groups reduces but does not eliminate selectivity relative to phosphatidylcholine. The dependence of the lipid selectivity on head group and chain composition is in the order cardiolipin approximately equal to monolysocardiolipin greater than acylcardiolipin greater than dimethylcardiolipin greater than phosphatidylcholine approximately equal to lysophosphatidylcholine, where acylcardiolipin has the spin-label chain attached at the center -OH of the head group. The degree of association of the negatively charged cardiolipin derivatives with cytochrome oxidase decreases with increasing salt concentration, to a level comparable to that for dimethylcardiolipin. At high ionic strength there is still a marked selectivity relative to phosphatidylcholine. Li+ ions are more effective in screening the interaction than are Na+ ions, and divalent ions are more effective than monovalent ions. The selectivity for cardiolipin is only slightly reduced on titrating the protein to high pH. Alkylation of the protein with N-ethylmaleimide has little effect on the titration behavior. Covalent modification of the protein by reaction with citraconic anhydride decreases the selectivity of interaction with cardiolipin. It is concluded that cardiolipin possesses an additional specificity of interaction with cytochrome oxidase other than that of purely electrostatic origin.
利用酰基链上自旋标记的脂质，通过电子自旋共振光谱研究了各种心磷脂类似物与牛肉心脏细胞色素氧化酶在与二肉豆蔻酰磷脂酰胆碱重构复合物中的相互作用选择性。在心磷脂及其单溶血衍生物之间未观察到选择性差异，同样，在磷脂酰胆碱和溶血磷脂酰胆碱之间也未观察到选择性。通过磷酸基团甲基化去除心磷脂电荷会降低但不会消除相对于磷脂酰胆碱的选择性。脂质选择性对头部基团和链组成的依赖性顺序为：心磷脂约等于单溶血心磷脂大于酰基心磷脂大于二甲基心磷脂大于磷脂酰胆碱约等于溶血磷脂酰胆碱，其中酰基心磷脂的自旋标记链连接在头部基团的中心 -OH 处。带负电荷的心磷脂衍生物与细胞色素氧化酶的结合程度随盐浓度增加而降低，降至与二甲基心磷脂相当的水平。在高离子强度下，相对于磷脂酰胆碱仍存在明显的选择性。Li⁺离子比 Na⁺离子更有效地屏蔽相互作用，二价离子比单价离子更有效。将蛋白质滴定至高 pH 时，对心磷脂的选择性仅略有降低。用 N - 乙基马来酰亚胺对蛋白质进行烷基化对滴定行为影响很小。通过与柠康酸酐反应对蛋白质进行共价修饰会降低与心磷脂相互作用的选择性。得出的结论是，心磷脂与细胞色素氧化酶的相互作用具有除纯静电起源之外的额外特异性。
