Rapid Hydrogen–Deuterium Exchange in Liquid Droplets

Abstract

The rate of hydrogen-deuterium exchange (HDX) in aqueous droplets of phenethylamine has been determined with submillisecond temporal resolution by mass spectrometry using nanoelectrospray ionization with a theta-capillary. The average speed of the microdroplets is measured using microparticle image velocimetry. The droplet travel time is varied from 20 to 320 μs by changing the distance between the emitter and the heated inlet to the mass spectrometer and the voltage applied to the emitter source. The droplets were found to accelerate by ∼30% during their observable travel time. Our droplet imaging shows that the theta-capillary produces two Taylor cone-jets (one per channel), causing mixing to take place from droplet fusion in the Taylor spray zone. Phenethylamine (ϕCHCHNH) was chosen to study because it has only one functional group (-NH) that undergoes rapid HDX. We model the HDX with a system of ordinary differential equations. The rate constant for the formation of -NHD from -NH is 3660 ± 290 s, and the rate constant for the formation of -NHD from -NHD is 3330 ± 270 s. The observed rates are about 3 times faster than what has been reported for rapidly exchangeable peptide side-chain groups in bulk measurements using stopped-flow kinetics and NMR spectroscopy. We also applied this technique to determine the HDX rates for a small 10-residue peptide, angiotensin I, in aqueous droplets, from which we found a 7-fold acceleration of HDX in the droplet compared to that in bulk solution.

Hydrogen/deuterium exchange in mass spectrometry

“…65,163 Recently the acceleration of the H/D exchange in microdroplets was demonstrated. 66,164 Authors estimated the velocity of droplets to be of the order ∼15 m/s before the entering the desolvating capillary.…”

“…The introduction of the atmospheric pressure ionization (API) sources opened the possibility to perform the H/D exchange reaction not in the high vacuum stage of the mass spectrometer but directly during the ionization at the atmospheric pressure, which showed several unexpected but favorable phenomena. For example, it was shown that chemical reactions (including H/D exchange) in charged microdroplets of electrospray are considerably accelerated in comparison with solutions . The first such experiments were performed by using thermospray and electrospray .…”

“…For example, it was shown that chemical reactions (including H/D exchange) in charged microdroplets of electrospray are considerably accelerated in comparison with solutions. [65][66][67] The first such experiments were performed by using thermospray 68 and electrospray. 69 Later the H/D exchange reaction in the course of ionization process was demonstrated by many other researchers for ESI, [70][71][72][73][74][75] APCI, 76 APPI, [77][78][79] and other types of ion sources.…”

Protein dynamics revealed by hydrogen/deuterium exchange mass spectrometry: Correlation between experiments and simulation

“…It should be noted that this HDX technique explores the protein conformations in the gas phase and its mechanism of exchange might be different from that of in‐solution HDX. More recently, fast HDX in liquid droplets determined by nano‐ESI‐MS using a theta‐capillary also realized sub‐millisecond HDX . The peptides and D 2 O from two channels of the capillary reacted during transit between the ESI emitter and MS inlet.…”

“…More recently, fast HDX in liquid droplets determined by nano-ESI-MS using a theta-capillary also realized sub-millisecond HDX. 37 The peptides and D 2 O from two channels of the capillary reacted during transit between the ESI emitter and MS inlet. Thus, the HDX could vary from ten to several hundred microseconds according to the configuration of the emitter and inlet.…”