Structure and Topology of the Huntingtin 1-17 Membrane Anchor by a Combined Solution and Solid-State NMR Approach
Structure and Topology of the Huntingtin 1-17 Membrane Anchor by a Combined Solution and Solid-State NMR Approach
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DOI：
10.1016/j.bpj.2013.06.030
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发表时间：
2013-08-06
影响因子：
3.4
通讯作者：
Bechinger, Burkhard
中科院分区：
文献类型：
--
作者：
Michalek, Matthias;Salnikov, Evgeniy S.;Bechinger, Burkhard
亨廷顿蛋白的氨基末端结构域直接位于该蛋白的多聚谷氨酰胺束的上游，在该大蛋白的几个重要性质和亨廷顿病的发展中起决定性作用。该亨廷顿蛋白1-17结构域一方面已知显著增加聚谷氨酰胺聚集速率，另一方面已显示参与细胞膜相互作用。在这里，我们确定了十二烷基磷酸胆碱胶束中的亨廷顿蛋白1-17的高分辨率结构和其螺旋结构域的拓扑结构在定向磷脂酰胆碱双层。使用二维溶液NMR光谱的低能量构象的多肽被确定在十二烷基磷酸胆碱洗涤剂胶束的存在下。在下一步中，从在选定位点用N-15和H-2标记的亨廷顿蛋白1-17获得一组四个固态NMR角约束。胶束合奏的螺旋构象只有一个有限的一套同意在定量的细节与固态角度限制亨廷顿蛋白1-17在支持的平面脂质双层。因此，固态核磁共振数据被用来进一步细化磷脂双层结构的域。同时，它的膜拓扑结构的确定和不同的运动机制的膜相关域进行了探索。亨廷顿蛋白1-17在膜缔合时的显著结构转变导致从K6到F17的α-螺旋构象，即，直到多聚谷氨酰胺束的最开始。这种两亲性螺旋几乎平行于膜表面排列（倾斜角类似于77度），其特征在于一侧的疏水脊和沿着螺旋亲水面的阳离子和阴离子残基的交替。这种排列有利于亨廷顿蛋白1-17结构域之间的静电相互作用，并可能与近端聚谷氨酰胺束。
The very amino-terminal domain of the huntingtin protein is directly located upstream of the protein's polyglutamine tract, plays a decisive role in several important properties of this large protein and in the development of Huntington's disease. This huntingtin 1-17 domain is on the one hand known to markedly increase polyglutamine aggregation rates and on the other hand has been shown to be involved in cellular membrane interactions. Here, we determined the high-resolution structure of huntingtin 1-17 in dodecyl phosphocholine micelles and the topology of its helical domain in oriented phosphatidylcholine bilayers. Using two-dimensional solution NMR spectroscopy the low-energy conformations of the polypeptide were identified in the presence of dodecyl phosphocholine detergent micelles. In a next step a set of four solid-state NMR angular restraints was obtained from huntingtin 1-17 labeled with N-15 and H-2 at selected sites. Of the micellar ensemble of helical conformations only a limited set agrees in quantitative detail with the solid-state angular restraints of huntingtin 1-17 obtained in supported planar lipid bilayers. Thereby, the solid-state NMR data were used to further refine the domain structure in phospholipid bilayers. At the same time its membrane topology was determined and different motional regimes of this membrane-associated domain were explored. The pronounced structural transitions of huntingtin 1-17 upon membrane-association result in a a-helical conformation from K6 to F17, i.e., up to the very start of the polyglutamine tract. This amphipathic helix is aligned nearly parallel to the membrane surface (tilt angle similar to 77 degrees) and is characterized by a hydrophobic ridge on one side and an alternation of cationic and anionic residues that run along the hydrophilic face of the helix. This arrangement facilitates electrostatic interactions between huntingtin 1-17 domains and possibly with the proximal polyglutamine tract.