Tamamizu-Kato Shiori, Cohen Jenny K, Drake Carolyn B, Kosaraju Malathi G, Drury Jessica, Narayanaswami Vasanthy
Center for the Prevention of Obesity, Cardiovascular Disease, and Diabetes, Children's Hospital Oakland Research Institute, 5700 Martin Luther King, Jr. Way, Oakland, California 94609, USA.
Biochemistry. 2008 May 6;47(18):5225-34. doi: 10.1021/bi702097s. Epub 2008 Apr 12.
Apolipoprotein (apo) E is an exchangeable apolipoprotein that plays an integral role in cholesterol transport in the plasma and the brain. It is also associated with protein misfolding or amyloid proteopathy of the beta amyloid peptide (Abeta) in Alzheimer's disease (AD) and cerebral amyloid angiopathy. The C-terminal domain (CT) of apoE encompasses two types of amphipathic alpha helices: a class A helix (residues 216-266) and a class G* helix (residues 273-299). This domain also harbors high-affinity lipoprotein binding and apoE self-association sites that possibly overlap. The objective of this study is to examine if the neurotoxic oligomeric Abeta interacts with apoE CT and if this association affects the lipoprotein binding function of recombinant human apoE CT. Site-specific fluorescence labeling of single cysteine-containing apoE CT variants with donor probes were employed to identify the binding of Abeta bearing an acceptor probe by intermolecular fluorescence resonance energy-transfer analysis. A higher efficiency of energy transfer was noted with probes located in the class A helix than with those located in the class G* helix of apoE CT. In addition, incubation of apoE CT with Abeta severely impaired the lipid binding ability and the overall amount of lipid-associated apoE CT. However, when apoE CT is present in a lipid-bound state, Abeta appears to be localized within the lipid milieu of the lipoprotein particle and not associated with any specific segments of the protein. When our data are taken together, they suggest that Abeta association compromises the fundamental lipoprotein binding function of apoE, which may have implications not only in terms of amyloid buildup but also in terms of the accumulation of cholesterol at extracellular sites.
载脂蛋白（apo）E是一种可交换的载脂蛋白，在血浆和大脑中的胆固醇转运中发挥着不可或缺的作用。它还与阿尔茨海默病（AD）中的β淀粉样肽（Aβ）的蛋白质错误折叠或淀粉样蛋白病以及脑淀粉样血管病有关。apoE的C末端结构域（CT）包含两种两亲性α螺旋：A类螺旋（残基216 - 266）和G类螺旋（残基273 - 299）。该结构域还含有可能重叠的高亲和力脂蛋白结合位点和apoE自缔合位点。本研究的目的是检查神经毒性寡聚Aβ是否与apoE CT相互作用，以及这种结合是否会影响重组人apoE CT的脂蛋白结合功能。通过分子间荧光共振能量转移分析，用供体探针对含单个半胱氨酸的apoE CT变体进行位点特异性荧光标记，以鉴定带有受体探针的Aβ的结合。与位于apoE CT的G类螺旋中的探针相比，位于A类螺旋中的探针观察到更高的能量转移效率。此外，apoE CT与Aβ孵育会严重损害脂质结合能力和脂质相关apoE CT的总量。然而，当apoE CT以脂质结合状态存在时，Aβ似乎定位于脂蛋白颗粒的脂质环境中，而不与蛋白质的任何特定片段相关。综合我们的数据表明，Aβ的结合损害了apoE的基本脂蛋白结合功能，这不仅可能与淀粉样蛋白堆积有关，还可能与细胞外部位胆固醇的积累有关。
