Electrospray ionization (ESI) of denatured proteins produces a mass spectrum with a broad distribution of multiply charged ions. Attaching fixed positive charges, specifically quaternary ammonium groups, to proteins at their carboxylic acid groups generates substantially higher charge states compared to the corresponding unmodified proteins in positive-mode ESI. Ion-ion reactions of these modified proteins with reagent anions leads to charge reduction by proton transfer. These proton transfer reactions cannot remove charge from the quaternary ammonium groups, which do not have a proton to transfer to the anion. Thus, one might expect charge reduction to stop at a single charge state equal to the number of fixed charges on the modified protein. However, ion-ion reactions yield charge states lower than this number of fixed charges due to anion attachment (adduction) to the proteins. Charge reduction via ion-molecule reactions involving gas-phase bases also give adducts on the modified protein ions in low charge states. Such adducts are avoided by keeping the ions in charge states well above the number of fixed charges. In the present work protein ions were selectively "parked" within an ion trap mass spectrometer in a high charge state by mild radiofrequency excitation that dramatically slows their ion-ion reaction rate-a technique termed "ion parking". The combination of ion parking with the fixed-charge modified proteins permits generation of a large population of ions in a single, very high charge state.

中文翻译：

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与固定电荷修饰蛋白质进行离子-离子反应，产生单一、非常高电荷态的离子


变性蛋白质的电喷雾电离 (ESI) 产生具有广泛分布的多电荷离子的质谱。与正模式 ESI 中相应的未修饰蛋白质相比，在蛋白质的羧酸基团上附加固定正电荷（特别是季铵基团）会产生明显更高的电荷态。这些修饰的蛋白质与试剂阴离子的离子反应通过质子转移导致电荷减少。这些质子转移反应不能去除季铵基团的电荷，季铵基团没有质子可转移到阴离子。因此，人们可能期望电荷减少停止在等于修饰蛋白质上固定电荷数量的单一电荷状态。然而，由于阴离子与蛋白质的附着（加合），离子-离子反应产生的电荷态低于该固定电荷数。通过涉及气相碱的离子-分子反应进行电荷还原，也会在低电荷态的修饰蛋白质离子上产生加合物。通过将离子保持在电荷状态远高于固定电荷的数量来避免这种加合物。在目前的工作中，通过温和的射频激发，蛋白质离子被选择性地“停泊”在离子阱质谱仪内，处于高电荷状态，这大大减慢了它们的离子-离子反应速率——一种称为“离子停泊”的技术。离子停泊与固定电荷修饰蛋白质的组合允许产生大量处于单一、非常高电荷状态的离子。