Electrosonic Spray Ionization. A Gentle Technique for Generating Folded Proteins and Protein Complexes in the Gas Phase and for Studying Ion−Molecule Reactions at Atmospheric Pressure

Abstract

Electrosonic spray ionization (ESSI), a variant on electrospray ionization (ESI), employs a traditional micro ESI source with supersonic nebulizing gas. The high linear velocity of the nebulizing gas provides efficient pneumatic spraying of the charged liquid sample. The variable electrostatic potential can be tuned to allow efficient and gentle ionization. This ionization method is successfully applied to aqueous solutions of various proteins at neutral pH, and its performance is compared to that of the nanospray and micro ESI techniques. Evidence for efficient desolvation during ESSI is provided by the fact that the peak widths for various multiply charged protein ions are an order of magnitude narrower than those for nanospray. Narrow charge-state distributions compared to other ESI techniques are observed also; for most of the proteins studied, more than 90% of the protein ions can be accumulated in one charge state using ESSI when optimizing conditions. The fact that the abundant charge state is normally as low or lower than that recorded by ESI or nanospray indicates that folded protein ions are generated. The sensitivity of the ionization technique to high salt concentrations is comparable to that of nanospray, but ESSI is considerably less sensitive to high concentrations of organic additives such as glycerol or 2-amino-2-(hydroxymethyl)-1,3-propanediol (Tris base). Noncovalent complexes are observed in the case of myoglobin, protein kinase A/ATP complex, and other proteins. The extent of dissociation of protein ions in ESSI is comparable to or even smaller than that in the case of nanospray, emphasizing the gentle nature of the method. The unique features of ESSI are ascribed to very efficient spraying and the low internal energy supplied to the ions. Evidence is provided that the method is capable of generating fully desolvated protein ions at atmospheric pressure. This allows the technique to be used for the study of ion-molecule reactions at atmospheric pressure and examples of this are shown.

Native Desorption Electrospray Ionization Liberates Soluble and Membrane Protein Complexes from Surfaces

“…We found that signal intensity was significantly improved if the length of the sample transfer tube, used in conventional DESI set‐ups,15 was minimised and the stage was located directly under the inlet of our ion source. We optimised signal intensity using hen egg‐white lysozyme and found that the spectra recorded under these native DESI conditions are similar to those from typical nanoflow ES capillaries, implying that the folded state of the protein is maintained (Figure 1 b).…”

Internal energy distributions in desorption electrospray ionization (DESI)

“…It should be noted though, that the initial events in the two experiments are quite different, and recent simulations show that the process of formation of progeny droplets from the initial ϳ4 m droplets in DESI is not dependent on charge build-up as it is in ESI. These dissimilarities need not result in differences in the final internal energy (E) since the later processes occurring in the atmospheric pressure interface are probably the same for these two processes and, as we will show later, dominate the energetics of the process.Here we use the "survival yield" (SY) method [42] applied to a set of thermometer ions (benzylpyridinium cations) to judge internal energy deposition associated with soft ionization techniques [43] to measure and compare the internal energy distributions of the ions generated by DESI and two related methods which produce ions directly from solution, namely ESI and electrosonic spray ionization (ESSI) [44]. We also examine the effect of various operating parameters in the DESI and ESSI sources on the extent of fragmentation of the thermometer ions.…”

“…Here we use the "survival yield" (SY) method [42] applied to a set of thermometer ions (benzylpyridinium cations) to judge internal energy deposition associated with soft ionization techniques [43] to measure and compare the internal energy distributions of the ions generated by DESI and two related methods which produce ions directly from solution, namely ESI and electrosonic spray ionization (ESSI) [44]. We also examine the effect of various operating parameters in the DESI and ESSI sources on the extent of fragmentation of the thermometer ions.…”

Characterization of acid-induced protein conformational changes and noncovalent complexes in solution by using coldspray ionization mass spectrometry

“…To preserve the native protein conformation, a significant amount of effort has also been made on modifications to the ESI source. The electrosonic spray ionization (ESSI) [31,32] method, using a traditional micro ESI source with supersonic nebulizing gas, allows for the characterization of protein complexes in solution. In addition, the ESI source combined with an air amplifier (ESI-AA) [33] also showed improvement in preserving folded conformations.…”