Wagner D S, Melton L G, Yan Y, Erickson B W, Anderegg R J
Glaxo Research Institute, Research Triangle Park, North Carolina 27709.
Protein Sci. 1994 Aug;3(8):1305-14. doi: 10.1002/pro.5560030817.
Deuterium exchange was monitored by electrospray ionization mass spectrometry (ESI-MS) to study the slowly exchanging (hydrogen bonded) peptide hydrogens of several alpha-helical peptides and beta-sheet proteins. Polypeptides were synthetically engineered to have mainly disordered, alpha-helical, or beta-sheet structure. For 3 isomeric 31-residue alpha-helical peptides, the number of slowly exchanging hydrogens as measured by ESI-MS in 50% CF3CD2OD (pD 9.5) provided estimates of their alpha-helicities (26%, 40%, 94%) that agreed well with the values (17%, 34%, 98%) measured by circular dichroic spectroscopy in the same nondeuterated solvent. For 3 betabellins containing a pair of beta-sheets and a related disordered peptide, their order of structural stability (12D > 12S > 14D > 14S) shown by their deuterium exchange rates in 10% CD3OD/0.5% CD3CO2D (pD 3.8) as measured by ESI-MS was the same as their order of structural stability to unfolding with increasing temperature or guanidinium chloride concentration as measured by circular dichroic spectroscopy in water. Compared to monitoring deuterium exchange by proton NMR spectrometry, monitoring deuterium exchange by ESI-MS requires much less sample (1-50 micrograms), much shorter analysis time (10-90 min), and no chemical quenching of the exchange reaction.
通过电喷雾电离质谱法（ESI-MS）监测氘交换，以研究几种α-螺旋肽和β-折叠蛋白中缓慢交换（氢键结合）的肽氢。对多肽进行合成改造，使其主要具有无序、α-螺旋或β-折叠结构。对于3种异构体的31个残基的α-螺旋肽，在50% CF3CD2OD（pD 9.5）中通过ESI-MS测量的缓慢交换氢的数量提供了它们α-螺旋度的估计值（26%、40%、94%），这与在相同非氘代溶剂中通过圆二色光谱法测量的值（17%、34%、98%）非常吻合。对于3种含有一对β-折叠的β-珠蛋白和一种相关的无序肽，通过ESI-MS在10% CD3OD/0.5% CD3CO2D（pD 3.8）中测量的氘交换率所显示的它们的结构稳定性顺序（12D > 12S > 14D > 14S）与通过圆二色光谱法在水中测量的它们随温度或氯化胍浓度升高而展开的结构稳定性顺序相同。与通过质子核磁共振光谱法监测氘交换相比，通过ESI-MS监测氘交换所需的样品量少得多（1 - 50微克），分析时间短得多（10 - 90分钟），并且无需对交换反应进行化学淬灭。
