Lala A K, Kaul P, Ratnam P B
Department of Chemistry, Indian Institute of Technology, Bombay, Powai, India.
J Protein Chem. 1995 Oct;14(7):601-9. doi: 10.1007/BF01886886.
The insertion of soluble proteins into membranes has been a topic of considerable interest. We have studied the insertion of bovine alpha-lactalbumin into single-bilayer vesicles prepared from egg phosphatidylcholine (PC). Fluorescence studies indicated rapid and tight binding of apo-alpha-lactalbumin (apo-alpha-LA) to PC vesicles as a function of pH. The binding was maximal at pH values which favor the formation of the molten globule state. As an increase of hydrophobic surface is observed in the molten globule state, this conformational state can provide a molecular basis for insertion of soluble proteins into membranes. The membrane-bound complex formed at low pH (3.0) could be isolated and was found to be stable at neutral pH. The structural characterization of the apo-alpha-LA-PC complex was studied by fluorescence quenching using iodide, acrylamide, and 9,10-dibromostearic acid. The results obtained indicated that some of the tryptophans of apo-alpha-LA were buried in the membrane interior and some were exposed on the outer side. Fluorescence quenching and CD studies indicated the membrane-bound conformation of apo-alpha-LA was some conformational state that is between the soluble, fully folded conformation and the molten globule state.
可溶性蛋白质插入膜中一直是一个备受关注的话题。我们研究了牛α-乳白蛋白插入由鸡蛋磷脂酰胆碱（PC）制备的单层囊泡中的情况。荧光研究表明，脱辅基α-乳白蛋白（apo-α-LA）与PC囊泡的结合迅速且紧密，这是pH的函数。在有利于形成熔球态的pH值下，结合作用最大。由于在熔球态中观察到疏水表面增加，这种构象状态可为可溶性蛋白质插入膜中提供分子基础。在低pH（3.0）下形成的膜结合复合物可以被分离出来，并且发现在中性pH下是稳定的。使用碘化物、丙烯酰胺和9,10-二溴硬脂酸通过荧光猝灭研究了apo-α-LA-PC复合物的结构特征。所得结果表明，apo-α-LA的一些色氨酸被埋在膜内部，一些则暴露在外侧。荧光猝灭和圆二色性研究表明，apo-α-LA的膜结合构象是介于可溶性、完全折叠构象和熔球态之间的某种构象状态。
