Wang G, Sparrow J T, Cushley R J
Institute of Molecular Biology and Biochemistry, Simon Fraser University, Burnaby, British Columbia, Canada V5A 1S6.
Biochemistry. 1997 Nov 4;36(44):13657-66. doi: 10.1021/bi971151q.
The conformation of a synthetic peptide of 46 residues from apoA-I was investigated by fluorescence, CD, and 2D NMR spectroscopies in lipid-mimetic environments. ApoA-I(142-187) is mainly unstructured in water but helical in SDS or dodecylphosphocholine (DPC), although the peptide only associates with DPC at approximately the critical micellar concentration. Solution structures of apoA-I(142-187) were determined by distance geometry calculations based on 450 (in DPC-d38) or 397 (in SDS-d25) NOE-derived distance restraints, respectively. Backbone RMSDs for superimposing the two helical regions 146-162 and 168-182 are 0.98 +/- 0.22 (2.38 +/- 0.20) and 1.99 +/- 0.42 (2.02 +/- 0.21) A in DPC (SDS), respectively. No interhelical NOE was found, suggesting that helix-helix interactions between the two helical domains in apoA-I(142-187) are unlikely. Similar average, curved helix-hinge-helix structures were found in both SDS and DPC micelles with the hydrophobic residues occupying the concave face, indicating that hydrophobic interactions dominate. Intermolecular NOESY experiments, performed in the presence of 50% protonated SDS, confirm that the two amphipathic helices and Y166 in the hinge all interact with the micelle. The involvement of Y166 in lipid binding is supported by fluorescence spectroscopy as well. On the basis of all the data above, we propose a model for the peptide-lipid complexes wherein the curved amphipathic helix-hinge-helix structural motif straddles the micelle. The peptide-aided signal assignment achieved for apoA-I(122-187) (66mer) and apoA-I suggests that such a structural motif is retained in the longer peptide and most likely in the intact protein.
在类脂环境中，通过荧光、圆二色光谱和二维核磁共振光谱研究了载脂蛋白A-I中一段含46个残基的合成肽的构象。载脂蛋白A-I(142 - 187)在水中主要呈无规结构，但在十二烷基硫酸钠（SDS）或十二烷基磷酰胆碱（DPC）中呈螺旋结构，尽管该肽仅在大约临界胶束浓度时与DPC结合。分别基于450个（在DPC-d38中）或397个（在SDS-d25中）由核Overhauser效应（NOE）得出的距离限制，通过距离几何计算确定了载脂蛋白A-I(142 - 187)的溶液结构。在DPC（SDS）中，将两个螺旋区域146 - 162和168 - 182进行叠加时，主链均方根偏差（RMSD）分别为0.98±0.22（2.38±0.20）和1.99±0.42（2.02±0.21）埃。未发现螺旋间的NOE，这表明载脂蛋白A-I(142 - 187)中两个螺旋结构域之间不太可能存在螺旋 - 螺旋相互作用。在SDS和DPC胶束中均发现了类似的平均弯曲螺旋 - 铰链 - 螺旋结构，疏水残基位于凹面，表明疏水相互作用起主导作用。在50%质子化SDS存在下进行的分子间NOESY实验证实，两个两亲性螺旋和铰链区的Y166均与胶束相互作用。荧光光谱也支持Y166参与脂质结合。基于上述所有数据，我们提出了一个肽 - 脂质复合物模型，其中弯曲的两亲性螺旋 - 铰链 - 螺旋结构基序横跨胶束。对载脂蛋白A-I(122 - 187)（66肽）和载脂蛋白A-I实现的肽辅助信号归属表明，这种结构基序在较长的肽中得以保留，并且很可能在完整蛋白质中也存在。
