Wilson Jesse W, Donor Micah T, Shepherd Samantha O, Prell James S
Department of Chemistry and Biochemistry, University of Oregon, 1253 University of Oregon, Eugene, Oregon 97403-1253, United States.
Materials Science Institute, University of Oregon, 1252 University of Oregon, Eugene, Oregon 97403-1252, United States.
J Am Soc Mass Spectrom. 2020 Jul 15. doi: 10.1021/jasms.0c00117.
Quadrupole ion mobility time-of-flight (Q-IM-TOF) mass spectrometers have revolutionized investigation of native biomolecular complexes. High pressures in the sources of these instruments aid transmission of protein complexes through damping of kinetic energy by collisional cooling. As adducts are removed through collisional heating (declustering), excessive collisional cooling can prevent removal of nonspecific adducts from protein ions, leading to inaccurate mass measurements, broad mass spectral peaks, and obfuscation of ligand binding. We show that reducing the source pressure using smaller aperture source sampling cones (SC) in a Waters Synapt G2-S instrument increases protein ion heating by decreasing collisional cooling, providing a simple way to enhance removal of adducted salts from soluble proteins (GroEL 14-mer) and detergents from a transmembrane protein complex (heptameric α-hemolysin, αHL). These experiments are supported by ion heating and cooling simulations which demonstrate reduced collisional cooling at lower source pressures. Using these easily swapped sample cones of different apertures is a facile approach to reproducibly extend the range of activation in Synapt-type instruments.
四极杆离子淌度飞行时间（Q-IM-TOF）质谱仪彻底改变了对天然生物分子复合物的研究。这些仪器源中的高压通过碰撞冷却来衰减动能，有助于蛋白质复合物的传输。由于加合物通过碰撞加热（去簇）被去除，过度的碰撞冷却会阻止非特异性加合物从蛋白质离子上被去除，从而导致质量测量不准确、质谱峰变宽以及配体结合情况模糊。我们表明，在沃特世Synapt G2-S仪器中使用较小孔径的源采样锥（SC）来降低源压力，通过减少碰撞冷却增加了蛋白质离子的加热，提供了一种简单的方法来增强从可溶性蛋白质（GroEL十四聚体）中去除加合盐以及从跨膜蛋白复合物（七聚体α-溶血素，αHL）中去除去污剂。这些实验得到了离子加热和冷却模拟的支持，模拟表明在较低源压力下碰撞冷却减少。使用这些易于更换的不同孔径的采样锥是一种简便的方法，可在Synapt型仪器中可重复地扩展激活范围。
