Huntingtin N-Terminal Monomeric and Multimeric Structures Destabilized by Covalent Modification of Heteroatomic Residues
Huntingtin N-Terminal Monomeric and Multimeric Structures Destabilized by Covalent Modification of Heteroatomic Residues
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DOI：
10.1021/acs.biochem.5b00478
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发表时间：
2015-07-21
期刊：
影响因子：
2.9
通讯作者：
Valentine, Stephen J.
中科院分区：
文献类型：
--
作者：
Arndt, James R.;Kondalaji, Samaneh Ghassabi;Valentine, Stephen J.
亨廷顿蛋白的早期低聚物形成可能是由蛋白中端（Nt17）的17个残基两亲性a-螺旋的自结合驱动的。寡聚结构与神经元毒性有关，可能是亨廷顿舞蹈病中重要的神经毒性物质。因此，Nt17的残基特异性结构表征对于理解和潜在抑制低聚物的形成至关重要。采用原生电喷雾离子迁移谱-质谱（IMS-MS）技术和分子动力学模拟（MDS）技术研究了独立于huntingtin外显子1剩余部分的Nt17共存的单体和多构象。MDS表明气相单体离子结构包括螺旋-旋转线圈结构和螺旋-延伸线圈区域。细长的二聚体包括部分螺旋的单体，排列成反平行的几何形状。这个堆叠的螺旋束可能代表了nt17驱动的低聚物形成的最早阶段。用焦碳酸二乙基（DEPC）进一步研究了Nt17单体和多聚体。一个n端位点（苏氨酸-3的n端）和赖氨酸-6在较高的DEPC浓度下被修饰，导致中间单体结构的形成。这些修饰导致延长的单体离子构象减少，以及多聚体形成的减少。从二聚体离子的MDS实验中，两种单体成分中的Lys6残基与邻近肽上的Ser16和Glul2残基相互作用；因此，多聚体形成的减少可能是由于这些或类似的相互作用的破坏。这项工作提供了一个结构选择模型，从中研究Nt17的自关联，并为Nt17的多聚化以及可能的亨廷顿蛋白外显子1聚集的早期阶段提供了重要的见解。
Early stage oligomer formation of the huntingtin protein may be driven by self-association of the 17-residue amphipathic a-helix at the protein's Nterminus (Nt17). Oligomeric structures have been implicated in neuronal toxicity and may represent important neurotoxic species in Huntington's disease. Therefore, a residue-specific structural characterization of Nt17 is crucial to understanding and potentially inhibiting oligomer formation. Native electrospray ion mobility spectrometry mass spectrometry (IMS-MS) techniques and molecular dynamics simulations (MDS) have been applied to study coexisting monomer and multirner conformations of Nt17, independent of the remainder of huntingtin exon 1. MDS suggests gas-phase monomer ion structures comprise a helix-turn-coil configuration and a helix-extended-coil region. Elongated dimer species comprise partially helical monomers arranged in an antiparallel geometry. This stacked helical bundle may represent the earliest stages of Nt17-driven oligomer formation. Nt17 monomers and multimers have been further probed using diethylpyrocarbonate (DEPC). An N-terminal site (N-terminus of Threonine-3) and Lysine-6 are modified at higher DEPC concentrations, which led to the formation of an intermediate monomer structure. These modifications resulted in decreased extended monomer ion conformers, as well as a reduction in multimer formation. From the MDS experiments for the dimer ions, Lys6 residues in both monomer constituents interact with Ser16 and Glul2 residues on adjacent peptides; therefore, the decrease in multimer formation could result from disruption of these or similar interactions. This work provides a structurally selective model from which to study Nt17 self-association and provides critical insight toward Nt17 multimerization and, possibly, the early stages of huntingtin exon 1 aggregation.