Stability of annexin V in ternary complexes with Ca2+ and anionic phospholipids: IR studies of monolayer and bulk phases.
Stability of annexin V in ternary complexes with Ca2+ and anionic phospholipids: IR studies of monolayer and bulk phases.
复制标题
膜联蛋白 V 在与 Ca2 和阴离子磷脂三元复合物中的稳定性：单层和体相的红外研究。
DOI：
10.1021/bi9819677
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发表时间：
1999
期刊：
影响因子：
2.9
通讯作者：
Mendelsohn,R
中科院分区：
文献类型：
--
作者：
Wu,F;Flach,CR;Seaton,BA;Mealy,TR;Mendelsohn,R
膜联蛋白V(Annexin V，AxV)是一类功能上依赖于钙离子与阴离子磷脂膜结合的蛋白质家族成员。用体相红外光谱(IR)和原位单分子膜反射−吸收光谱(IRRAS)研究了蛋白质的结构和稳定性随钙离子和磷脂的变化。体相实验表明，AxV在∼45−50°C时发生了不可逆热变性，在1617和1682 cm-1处出现了酰胺I带。然而，即使在60°C时，仍保留了一些天然的二级结构，与部分展开的“熔融球状”状态一致。与AxV单独、Ca~(2+)/AxV或脂质/AxV混合物相比，形成的Ca~(2+)/磷脂/蛋白质三元复合体显著保护蛋白质免受热变性。IRRAS还观察到在钙离子存在的情况下DMPA单层对AxV二级结构的稳定作用。在钙离子存在或不存在的情况下，吸附的AxV膜的光谱显示酰胺I模式发生了10 cm-1的位移，对应于A/W界面的有序结构的丧失。在体相和红外光谱实验中，仅在三元络合物中增强了对AxV中H→D交换的保护。结合数据表明，AxV的二级结构受到三元复合体中钙离子/膜组分的强烈影响，而脂类构象顺序不受蛋白质的影响。
Annexin V (AxV) is a member of a family of proteins that exhibit functionally relevant Ca2+-dependent binding to anionic phospholipid membranes. Protein structure and stability as a function of Ca2+and phospholipids was studied by bulk phase infrared (IR) spectroscopy and by IR reflection−absorption spectroscopy (IRRAS) of monolayers in situ at the air/water (A/W) interface. Bulk phase experiments revealed that AxV undergoes an irreversible thermal denaturation at ∼45−50 °C, as shown by the appearance of amide I bands at 1617 and 1682 cm-1. However, some native secondary structure is retained, even at 60 °C, consistent with a partially unfolded “molten globule” state. Formation of the Ca2+/phospholipid/protein ternary complex significantly protects the protein from thermal denaturation as compared to AxV alone, Ca2+/AxV, or lipid/AxV mixtures. Stabilization of AxV secondary structure by a DMPA monolayer in the presence of Ca2+was also observed by IRRAS. Spectra of an adsorbed AxV film in the presence or absence of Ca2+showed a 10 cm-1shift in the amide I mode, corresponding to loss of ordered structure at the A/W interface. In both the bulk phase and IRRAS experiments, protection against H→D exchange in AxV was enhanced only in the ternary complex. The combined data suggest that the secondary structure of AxV is strongly affected by the Ca2+/membrane component of the ternary complex whereas lipid conformational order is unchanged by protein.