Single homopolypeptide chains collapse into mechanically rigid conformations
Single homopolypeptide chains collapse into mechanically rigid conformations
复制标题
DOI：
10.1073/pnas.0900678106
复制
发表时间：
2009-08-04
影响因子：
11.1
通讯作者：
Fernandez, Julio M.
中科院分区：
文献类型：
--
作者：
Dougan, Lorna;Li, Jingyuan;Fernandez, Julio M.
亨廷顿氏病与谷氨酰胺（Q）插入蛋白亨廷顿蛋白中有关，导致多聚谷氨酰胺（polyQ）扩增，其自缔合形成聚集体。虽然polyQ聚集一直是深入研究的主题，但缺乏对单个polyQ链的相应透彻理解。在这里，我们展示了一种单分子力钳技术，直接探测单polyQ链的机械性能。我们已经制备了不同长度的polyQ构建体，其跨越正常和患病polyQ扩增的长度范围。每个polyQ构建体的侧翼是I27肌联蛋白模块，提供了被拉分子的清晰机械指纹。值得注意的是，在施加力的情况下，没有观察到任何polyQ构建体的延伸。这与肌联蛋白的无规卷曲蛋白PEVK形成直接对比，后者在力的作用下容易延伸。我们的测量表明，polyQ链形成机械稳定的塌陷结构。我们通过插入脯氨酸残基破坏polyQ链来测试这一假设，并发现它们的机械延伸性对脯氨酸中断的位置敏感。这些实验表明，polyQ链塌缩形成具有机械弹性的异构构象系综。我们进一步使用热退火分子动力学协议，广泛搜索构象空间，发现polyQ可以存在于高度机械稳定的紧凑的球状构象。这些塌陷结构的机械刚性可能超过真核蛋白酶体的功能能力，导致未消化的polyQ序列在体内积累。化学
Huntington's disease is linked to the insertion of glutamine (Q) in the protein huntingtin, resulting in polyglutamine (polyQ) expansions that self-associate to form aggregates. While polyQ aggregation has been the subject of intense study, a correspondingly thorough understanding of individual polyQ chains is lacking. Here we demonstrate a single molecule force-clamp technique that directly probes the mechanical properties of single polyQ chains. We have made polyQ constructs of varying lengths that span the length range of normal and diseased polyQ expansions. Each polyQ construct is flanked by the I27 titin module, providing a clear mechanical fingerprint of the molecule being pulled. Remarkably, under the application of force, no extension is observed for any of the polyQ constructs. This is in direct contrast with the random coil protein PEVK of titin, which readily extends under force. Our measurements suggest that polyQ chains form mechanically stable collapsed structures. We test this hypothesis by disrupting polyQ chains with insertions of proline residues and find that their mechanical extensibility is sensitive to the position of the proline interruption. These experiments demonstrate that polyQ chains collapse to form a heterogeneous ensemble of conformations that are mechanically resilient. We further use a heat-annealing molecular dynamics protocol to extensively search the conformation space and find that polyQ can exist in highly mechanically stable compact globular conformations. The mechanical rigidity of these collapsed structures may exceed the functional ability of eukaryotic proteasomes, resulting in the accumulation of undigested polyQ sequences in vivo. CHEMISTRY