Apolipoprotein A-I assumes a "looped belt" conformation on reconstituted high density lipoprotein
Apolipoprotein A-I assumes a "looped belt" conformation on reconstituted high density lipoprotein
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DOI：
10.1074/jbc.m602077200
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发表时间：
2006-07-21
影响因子：
4.8
通讯作者：
Oda, Michael N.
中科院分区：
文献类型：
--
作者：
Martin, Dale D. O.;Budamagunta, Madhu S.;Oda, Michael N.
载脂蛋白A-I（apoA-I）在胆固醇逆向转运途径中起着重要作用，然而，其抗动脉粥样硬化作用的结构基础仍然知之甚少。在这里，我们采用EPR光谱和荧光共振能量转移来阐明盘状（9.4 nm）重构高密度脂蛋白（rHDL）的apoA-I单体的构象和相对对齐。EPR光谱提供了证据的扩展螺旋二级结构。位置139，因为它是检查的唯一显示与柔性环结构一致的动态运动特征的残基。在位置133和146处的氮氧探针的EPR谱表现出自旋耦合，表明这些位置接近于rHDL周边上的apoA-I配对对应物。使用具有单个色氨酸（能量供体）和/或单个半胱氨酸（其巯基部分用外部能量受体共价标记）的工程化apoA-I变体的荧光共振能量转移研究提供了证据，即围绕rHDL周边的成对apoA-I分子以延伸的反平行构象排列。结合位于插入序列位置（134 - 145）的氮氧探针的EPR谱不显示自旋耦合的观察，这导致我们提出了“环带”模型，其中残基133 - 146包含赋予apoA-I适应其结构以适应变化的颗粒脂质含量的内在能力的柔性环段。具体而言，在环带模型中，除了氨基酸134 - 145之外，apoA-I在螺旋5-螺旋5注册表中与其对应物对齐，以位置139为中心。
Apolipoprotein A-I(apoA-I) plays a central role in the reverse cholesterol transport pathway; however, the structural basis for its antiatherogenic effects remains poorly understood. Here we employ EPR spectroscopy and fluorescence resonance energy transfer to elucidate the conformation and relative alignment of apoA-I monomers on discoidal (9.4 nm) reconstituted high density lipoprotein (rHDL). EPR spectroscopy provided evidence for an extended helical secondary structure. Position 139 since it was the only residue examined to display a dynamic motional character consistent with a flexible loop structure. The EPR spectra of nitroxide probes at positions 133 and 146 exhibit spin coupling, indicating that these positions are proximal to an apoA-I paired counterpart on the perimeter of rHDL. fluorescence resonance energy transfer studies employing engineered apoA-I variants possessing a single tryptophan (energy donor) and/or a single cysteine (whose thiol moiety was covalently labeled with an extrinsic energy acceptor) provided evidence that paired apoA-I molecules around the perimeter of rHDL align in an extended antiparallel conformation. Taken together with the observation that the EPR spectra of nitroxide probes positioned at intervening sequence positions (134 - 145) do not exhibit spin coupling, this has led us to propose a "looped belt" model, wherein residues 133 - 146 comprise a flexible loop segment that confers to apoA-I an intrinsic ability to adapt its structure to accommodate changing particle lipid content. Specifically, in the looped belt model, with the exception of amino acids 134 - 145, apoA-I aligns with its counterpart in a helix 5-helix 5 registry, centered at position 139.