Dehydration of the lipid-protein microinterface on binding of phospholipase A2 to lipid bilayers.
Dehydration of the lipid-protein microinterface on binding of phospholipase A2 to lipid bilayers.
复制标题
磷脂酶 A2 与脂质双层结合时脂质-蛋白质微界面的脱水。
DOI：
10.1016/0005-2736(87)90002-2
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发表时间：
1987
期刊：
影响因子：
--
通讯作者：
Vaz,WL
中科院分区：
文献类型：
--
作者：
Jain,MK;Vaz,WL
描述了一种新方法来证明猪胰磷脂酶 A2 和该蛋白质结合的脂质双层之间的微界面的大量水相是不可接近的。该方法基于以下事实：与在氘化水中相比，蛋白质的色氨酸残基和连接至脂质的5-二甲氨基萘-1-磺酰基（丹酰基）发色团在水中的荧光发射量子产率较低。这些发色团的荧光发射量子产率是在蛋白质结合或不结合含有丹磺酰发色团的脂质双层表面的条件下在水和氘水中测量的。在蛋白质与双层表面紧密结合的条件下，两种荧光团的去溶剂化消除了观察到的氘化水的效果。结合的磷脂酶 A2 中的色氨酸残基也变得无法被丙烯酰胺或琥珀酰亚胺猝灭。仅在对磷脂酶 A2 的催化作用具有重要意义的条件下，才能观察到微界面的去溶剂化，而在跳跃模式下则不然。此外，在类似条件下，磷脂酶原 A2 与阴离子囊泡的结合不会导致微界面脱水。讨论了这些观察结果对于脂质-蛋白质相互作用的机械意义，特别是对于界面催化和界面活化的意义。
A novel method is described to demonstrate inaccessibility to the bulk aqueous phase of the microinterface between pig pancreatic phospholipase A2and lipid bilayers to which this protein is bound. The method is based on the fact that the fluorescence emission quantum yields of the tryptophan residue of the protein and of a 5-dimethylaminonaphthalene-1-sulfonyl (dansyl) chromophore attached to a lipid are lower in water as compared to that in deuterated water. The fluorescence emission quantum yield of these chromophores is measured in water and in deuterated water under conditions where the protein is either bound or not bound to the surface of a lipid bilayer containing the dansyl chromophore. Under conditions where the protein is tightly bound to the surface of the bilayer, desolvation of both fluorophores abolishes the observed effect of deuterated water. The tryptophan residue in the bound phospholipase A2also becomes inaccessible to fluorescence quenching by acrylamide or succinimide. Desolvation of the microinterface is observed only under conditions that are significant for the catalytic action of phospholipase A2in the scooting mode and not in the hopping mode. Also, under similar conditions, binding of pro-phospholipase A2to anionic vesicles does not cause dehydration of the microinterface. The mechanistic significance of these observations for lipid-protein interactions, in general, and for interfacial catalysis and interfacial activation, in particular, is discussed.