Structural and dynamic properties of juxta-membrane segments of caveolin-1 and caveolin-2 at the membrane interface
Structural and dynamic properties of juxta-membrane segments of caveolin-1 and caveolin-2 at the membrane interface
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DOI：
10.1007/s00249-009-0548-4
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发表时间：
2010-01-01
影响因子：
2
通讯作者：
Jamin, Nadege
中科院分区：
文献类型：
--
作者：
Le Lan, Charlotte;Gallay, Jacques;Jamin, Nadege
窖蛋白（caveolins，cav 1 -3）是细胞膜上的一种重要蛋白。cav-1的小窝蛋白支架结构域包括在其C-末端含有CRAC基序（V94 TKYWFYR 101）的短序列。为了在原子水平上研究这个基序在小窝蛋白-膜相互作用中的作用，我们对cav-1（V94-L102）九肽进行了详细的结构和动力学表征，该九肽包含这个基序，并包括cav-1疏水结构域（L102）的第一个残基，在十二烷基麦芽糖苷（DM）或十二烷基磷酸胆碱（DPC）胶束中，作为膜模拟物。荧光滴定和圆二色谱结果表明，Cav-1（V94-L102）在DPC和DM/阴离子脂质胶束中的分配比在DM胶束中的分配好。NMR数据显示，该肽折叠为位于DPC胶束的极性头基区域的两亲性螺旋。两侧为精氨酸和赖氨酸残基的两条酪氨酸侧链位于该螺旋的一面，而苯丙氨酸和色氨酸侧链位于相对面。荧光研究表明显着的色氨酸subnanosecond旋转，几个旋转异构体的存在下，和水/胶束界面内的异质位置。较短的cav-1（V94-R101）肽和cav-2（I79 SKYVMYKF 87）的同源序列的NMR研究允许描述L102和cav-2中发生的氨基酸变异对DPC胶束中的结构和定位的影响。基于小窝蛋白的拓扑模型，我们的研究结果表明，cav-1和cav-2九肽研究形成界面α-螺旋膜锚定，其中K/RhhhYK/Rh基序，也发现在cav-3，可能发挥重要作用。
Caveolins (cav1-3) are essential membrane proteins found in caveolae. The caveolin scaffolding domain of cav-1 includes a short sequence containing a CRAC motif (V94TKYWFYR101) at its C-terminal end. To investigate the role of this motif in the caveolin-membrane interaction at the atomic level, we performed a detailed structural and dynamics characterization of a cav-1(V94-L102) nonapeptide encompassing this motif and including the first residue of cav-1 hydrophobic domain (L102), in dodecylmaltoside (DM) or dodecylphosphocholine (DPC) micelles, as membrane mimics. Cav-1(V94-L102) partitioned better in DPC and in DM/anionic lipid micelles than in DM micelles, as shown by fluorescence titration and CD. NMR data revealed that this peptide folded as an amphipathic helix located in the polar head group region of DPC micelles. The two tyrosine side-chains, flanked by arginine and lysine residues, are situated on one face of this helix, whereas the phenylalanine and tryptophan side-chains are located on the opposite face. Fluorescence studies showed significant Trp subnanosecond rotations, the presence of several rotamers, and a heterogeneous location within the water/micelle interface. NMR studies of the shorter cav-1(V94-R101) peptide and of the homologous sequence of cav-2(I79SKYVMYKF87) allowed the description of the effect of L102 and of the amino acid variations occurring in cav-2 on the structure and localization in DPC micelles. Based on the topological model of caveolins, our results suggest that the cav-1 and cav-2 nonapeptides studied form interfacial alpha-helix membrane anchors in which the K/RhhhYK/Rh motif, also found in cav-3, may play a significant role.