Department of Chemistry, Technical University of Darmstadt, 64287 Darmstadt, Germany.
School of Engineering, University of Warwick, Coventry CV4 7AL, U.K.
J Am Soc Mass Spectrom. 2021 Jan 6;32(1):364-372. doi: 10.1021/jasms.0c00373. Epub 2020 Nov 25.
One of the main characteristics of biomolecular ions in mass spectrometry is their net charge, and a range of approaches exist to either increase or decrease this quantity in the gas phase. In the context of small molecules, it is well known that, in addition to the charge state, the charge site also has a profound effect on an ion's gas-phase behavior; however, this effect has been far less explored for peptides and intact proteins. Methods exist to determine charge sites of protein ions, and others have observed that the interplay of electrostatic repulsion and inherent basicity leads to different sites gaining or losing a charge depending on the total net charge. Here, we report two distinct protonation site isomers ("protomers") of α-synuclein occurring at the same charge state. The protomers showed important differences in their gas-phase fragmentation behavior and were furthermore distinguishable by ion mobility spectrometry. One protomer was produced under standard electrospray conditions, while the other was observed after addition of 10% dimethyl sulfoxide to the protein solution. Charge sites for both protomers were determined using ultraviolet photodissociation.
生物分子离子在质谱中的一个主要特征是其净电荷，并且存在多种方法可以在气相中增加或减少这种数量。在小分子的情况下，人们已经知道，除了电荷状态外，电荷位置对离子的气相行为也有深远的影响；然而，对于肽和完整蛋白质，这种影响的研究要少得多。已经存在确定蛋白质离子电荷位置的方法，并且其他人已经观察到静电排斥和固有碱性之间的相互作用导致不同的位置根据总净电荷获得或失去电荷。在这里，我们报告了α-突触核蛋白在相同电荷状态下发生的两种不同的质子化位置异构体（“前体”）。前体在其气相碎裂行为方面表现出重要差异，并且通过离子淌度谱法可以区分。一种前体是在标准电喷雾条件下产生的，而另一种是在向蛋白质溶液中添加 10%二甲基亚砜后观察到的。使用紫外线光解确定了两种前体的电荷位置。
