Arndt James R, Brown Robert J, Burke Kathleen A, Legleiter Justin, Valentine Stephen J
C. Eugene Bennett Department of Chemistry, West Virginia University, Morgantown, WV, 26506, USA.
J Mass Spectrom. 2015 Jan;50(1):117-26. doi: 10.1002/jms.3504.
Huntington's disease is a genetic neurodegenerative disorder caused by an expansion in a polyglutamine domain near the N-terminus of the huntingtin (htt) protein that results in the formation of protein aggregates. Here, htt aggregate structure has been examined using hydrogen-deuterium exchange techniques coupled with tandem mass spectrometry. The focus of the study is on the 17-residue N-terminal flanking region of the peptide that has been shown to alter htt aggregation kinetics and morphology. A top-down sequencing strategy employing electron transfer dissociation is utilized to determine the location of accessible and protected hydrogens. In these experiments, peptides aggregate in a deuterium-rich solvent at neutral pH and are subsequently subjected to deuterium-hydrogen back-exchange followed by rapid quenching, disaggregation, and tandem mass spectrometry analysis. Electrospray ionization of the peptide solution produces the M + 5H to M + 10H charge states and reveals the presence of multiple peptide sequences differing by single glutamine residues. The M + 7H to M + 9 charge states corresponding to the full peptide are used in the electron transfer dissociation analyses. Evidence for protected residues is observed in the 17-residue N-terminal tract and specifically points to lysine residues as potentially playing a significant role in htt aggregation.
亨廷顿舞蹈症是一种遗传性神经退行性疾病，由亨廷顿蛋白（htt）N端附近的多聚谷氨酰胺结构域扩张引起，导致蛋白质聚集体形成。在此，已使用氢-氘交换技术结合串联质谱对htt聚集体结构进行了研究。该研究的重点是肽段的17个残基N端侧翼区域，该区域已被证明会改变htt的聚集动力学和形态。采用电子转移解离的自上而下测序策略用于确定可及氢和受保护氢的位置。在这些实验中，肽段在中性pH的富氘溶剂中聚集，随后进行氘-氢反向交换，接着快速淬灭、解聚和串联质谱分析。肽溶液的电喷雾电离产生M + 5H至M + 10H电荷态，并揭示了存在多个相差单个谷氨酰胺残基的肽序列。对应于完整肽段的M + 7H至M + 9电荷态用于电子转移解离分析。在17个残基的N端区域观察到受保护残基的证据，特别指出赖氨酸残基可能在htt聚集中起重要作用。
