Wright P John, Zhang Jianmin, Douglas D J
Department of Chemistry, University of British Columbia, Vancouver, British Columbia, Canada.
J Am Soc Mass Spectrom. 2008 Dec;19(12):1906-13. doi: 10.1016/j.jasms.2008.07.018. Epub 2008 Jul 24.
At low pH in solutions of 50% methanol, proteins form expanded denatured states (the "H" state). In 90% methanol, proteins form expanded helical denatured states with artificial alpha-helices (the "H(c)" state). Gas-phase ions of ubiquitin, cytochrome c, apomyoglobin, and native and disulfide-reduced beta-lactoglobulin were formed by electrospray ionization (ESI) of the proteins from the H and H(c) states in solution. Both states in solution produce the same charge states in ESI. The conformations of the ions were studied with cross section measurements and gas-phase H/D exchange experiments. The cross sections show that the ions retain considerable folded structure. For a given protein and given charge state, ions produced from the H and H(c) states showed the same cross sections (within approximately 1%). Ions of cytochrome c, apomyoglobin, and native and reduced beta-lactoglobulin of a given charge state showed no differences in H/D exchange level when produced from the H or H(c) state. However, ubiquitin ions produced from the H(c) state consistently exchange fewer ( approximately 13%) hydrogens than ions produced from the H state, suggesting that in this case the gas-phase protein ions retain some memory of their solution conformations.
在50%甲醇溶液的低pH值条件下，蛋白质会形成伸展的变性状态（“H”状态）。在90%甲醇中，蛋白质会形成带有人工α螺旋的伸展螺旋变性状态（“H(c)”状态）。通过对溶液中处于H和H(c)状态的蛋白质进行电喷雾电离（ESI），形成了泛素、细胞色素c、脱辅基肌红蛋白以及天然和二硫键还原的β-乳球蛋白的气相离子。溶液中的这两种状态在电喷雾电离中产生相同的电荷状态。通过截面测量和气相H/D交换实验研究了离子的构象。截面显示离子保留了相当多的折叠结构。对于给定的蛋白质和给定的电荷状态，由H和H(c)状态产生的离子显示出相同的截面（误差约为1%）。对于给定电荷状态的细胞色素c、脱辅基肌红蛋白以及天然和还原的β-乳球蛋白离子，当由H或H(c)状态产生时，在H/D交换水平上没有差异。然而，由H(c)状态产生的泛素离子始终比由H状态产生的离子少交换约13%的氢，这表明在这种情况下，气相蛋白质离子保留了其溶液构象的一些记忆。
