Grishchenko V M, Kalinichenko L P, Deikus G Y, Veprintsev D B, Cawthern K M, Berliner L J, Permyakov E A
Institute of Theoretical and Experimental Biophysics, Russian Academy of Sciences, Pushchino, Moscow region, Russia.
Biochem Mol Biol Int. 1996 Mar;38(3):453-66.
Complexes of alpha-lactalbumin (alpha-LA)1 with dimyristoylphosphatidylcholine (DMPC) or dipalmitoylphosphatidylcholine (DPPC) liposomes at pH 8 and at pH 2 have been obtained by means of gel filtration. Thermal denaturation of alpha-LA complexes of DMPC or DPPC at pH 8 was found to depend on the saturation of protein by metal cations. The intrinsic fluorescence of DMPC-alpha-LA and DPPC-alpha-LA was sensitive to two thermal transitions. The first transition corresponded to the Tc of the lipid vesicles, while the second transition arose from the denaturation of the protein. Fluorescence spectrum position suggested that at low temperature tryptophan accessibility increases upon protein-DMPC or protein-DPPC association. At temperatures above the protein transition (70 degrees C) tryptophan appears to interact significantly with the apolar phase of DMPC and DPPC, evidenced by spectral blue shifts. Whereas the free protein at pH 2 adopts the molten globule (MG) state and is characterized by the absence of a thermal transition, the rapidly-isolated DMPC-alpha-LA complex was characterized by the appearance of a distinct fluorescence thermal transition between 50 and 60 degrees C. This result is consistent with a model of a partially-inserted form of alpha-LA which may possess some degree of tertiary structure and therefore unfolds cooperatively.
通过凝胶过滤法获得了α-乳白蛋白（α-LA）与二肉豆蔻酰磷脂酰胆碱（DMPC）或二棕榈酰磷脂酰胆碱（DPPC）脂质体在pH 8和pH 2条件下的复合物。发现在pH 8时，DMPC或DPPC的α-LA复合物的热变性取决于金属阳离子对蛋白质的饱和程度。DMPC-α-LA和DPPC-α-LA的内在荧光对两个热转变敏感。第一个转变对应于脂质囊泡的Tc，而第二个转变源于蛋白质的变性。荧光光谱位置表明，在低温下，色氨酸的可及性在蛋白质与DMPC或蛋白质与DPPC结合时增加。在高于蛋白质转变温度（70℃）时，色氨酸似乎与DMPC和DPPC的非极性相发生显著相互作用，光谱蓝移证明了这一点。在pH 2时，游离蛋白质呈熔融球状（MG）状态，其特征是没有热转变，而快速分离的DMPC-α-LA复合物的特征是在50至60℃之间出现明显的荧光热转变。这一结果与α-LA部分插入形式的模型一致，该模型可能具有一定程度的三级结构，因此会协同展开。
