Determination of gas phase protein ion densities via ion mobility analysis with charge reduction

Abstract

We use a charge reduction electrospray (ESI) source and subsequent ion mobility analysis with a differential mobility analyzer (DMA, with detection via both a Faraday cage electrometer and a condensation particle counter) to infer the densities of single and multiprotein ions of cytochrome C, lysozyme, myoglobin, ovalbumin, and bovine serum albumin produced from non-denaturing (20 mM aqueous ammonium acetate) and denaturing (1 : 49.5 : 49.5, formic acid : methanol : water) ESI. Charge reduction is achieved through use of a Po-210 radioactive source, which generates roughly equal concentrations of positive and negative ions. Ions produced by the source collide with and reduce the charge on ESI generated drops, preventing Coulombic fissions, and unlike typical protein ESI, leading to gas-phase protein ions with +1 to +3 excess charges. Therefore, charge reduction serves to effectively mitigate any role that Coulombic stretching may play on the structure of the gas phase ions. Density inference is made via determination of the mobility diameter, and correspondingly the spherical equivalent protein volume. Through this approach it is found that for both non-denaturing and denaturing ESI-generated ions, gas-phase protein ions are relatively compact, with average densities of 0.97 g cm(-3) and 0.86 g cm(-3), respectively. Ions from non-denaturing ESI are found to be slightly more compact than predicted from the protein crystal structures, suggesting that low charge state protein ions in the gas phase are slightly denser than their solution conformations. While a slight difference is detected between the ions produced with non-denaturing and denaturing ESI, the denatured ions are found to be much more dense than those examined previously by drift tube mobility analysis, in which charge reduction was not employed. This indicates that Coulombic stretching is typically what leads to non-compact ions in the gas-phase, and suggests that for gas phase measurements to be correlated to biomolecular structures in solution, low charge state ions should be analyzed. Further, to determine if different solution conditions give rise to ions of different structure, ions of similar charge state should be compared. Non-denatured protein ion densities are found to be in excellent agreement with non-denatured protein ion densities inferred from prior DMA and drift tube measurements made without charge reduction (all ions with densities in the 0.85-1.10 g cm(-3) range), showing that these ions are not strongly influenced by Coulombic stretching nor by analysis method.

Collision cross section calculations for polyatomic ions considering rotating diatomic/linear gas molecules

“…These ions are selected for examination because not only they have been well characterized by IMS-MS measurements, 31, 32, 39-42 but also they are either sufficiently small to generate candidate structures in the gas phase (tetraalkylammonium ions) 9 or have clear, charge dependent structures in the gas phase (PEG ions). 32,40 We note this is in contrast to often studied protein and multiprotein complex ions, which do not necessarily have structures resembling their crystal structures or those observed in solution upon introduction in the gas phase; [43][44][45][46][47][48][49] the examination of protein ions brings an extra degree of ambiguity into comparison between different collisionreemission models and measurements. Figure 4 displays a comparison of the measured 31 inverse mobilities in air at atmospheric pressure at 293 K for tetramethylammonium + (TMA + ), tetrapropylammonium + (TPA + ), tetrabutylammonium + (TBA + ), tetraheptylammonium + (THA + ), tetradecylammonium + (TDA + ), and tetradodecylammonium + (TDDA + ) ions to model predictions for all 4 cases and with both model D and model M gas molecules.…”

Electrospray–differential mobility analysis of bionanoparticles

“…This work was followed over the next decade by several reports from Allmaier et al [11], Wick et al [12] and de la Mora et al [13] related to characterization of several other proteins, viruses and polymers. Starting in 2006 as seen in Figures 1.3A and 1.3B a dramatic increase in the number of publications and citations related to ES-DMA occurred with contributions from Biswas et al [14], Zachariah et al [15], Loo et al [16], Hogan et al [17], Pergantis et al [18], Pease et al [19] and Hackley et al [20]. Although developed primarily for size determination, with an increasing pool of users, several orthogonal utilities have been found for this technique a detailed discussion of which will be appear later in Chapter 3.…”

Atomic Cluster Generation with an Atmospheric Pressure Spark Discharge Generator

“…While DMAs have been designed originally for particles in the size range of ten to several hundreds of nanometers (Knutson and Whitby 1975;Reischl 1991;Chen et al 1996), the DMA used in this study (half-mini DMA, NanoEngineering, Inc.) has been designed to enable mobility measurements of clusters and ions down to sub-nanometer size ranges (De Juan and Fern andez de la Mora 1998; Maißer et al 2011;Attoui et al 2013;de la Mora and Kozlowski 2013;Wang et al 2014). The design of the half-mini DMA allows use of sheath flow rates of several hundreds of liters per minute that get sonic at values >700 lpm for air.…”