Abstract

We present a new variable temperature (VT), high resolution ion mobility (IM) drift tube coupled to a commercial mass spectrometer (MS). Ions are generated in an electrospray ion source with a sampling cone interface and two stacked ring RF guides which transfer ions into the mobility analyzer located prior to a quadrupole time-of-flight mass spectrometer. The drift cell can be operated over a pressure range of 0.5-3 Torr and a temperature range of 150-520 K with applied fields typically between 3 and 14 V cm-1. This makes the instrument suitable for rotationally averaged collision cross section (CCS) measurements at low E/N ratios where ions are near thermal equilibrium with the buffer gas. Fundamental studies of the effective ion temperatures can be performed at high E/N ratios. An RF ion trap/buncher is located at the beginning of the drift region, which modulates the continuous ion beam into spatially narrow packets. Packets of ions then drift in a linear electric field, which is 50.5 cm long, and are separated according to their mobility in an inert buffer gas. Post-drift, an ion funnel focuses the radially spread pulses of ions into the inlet of a commercial MS platform (Micromass QToF2). We present the novel features of this instrument and results from VT-IM-MS experiments on a range of model systems-IMS CCS standards (Agilent ESI Tune Mix), the monomeric protein Ubiquitin (8.6 kDa), and the tetrameric protein complex Concanavalin A (103 kDa). We evaluate the performance of the instrument by comparing ambient DTCCSHe values of model compounds with those found in the literature. Several effects of temperature on collision cross sections and resolution are observed. For small rigid molecules, changes in resolution are consistent with anticipated thermal diffusion effects. Changes in measured DTCCSHe for these rigid systems at different temperatures are attributed primarily to the effect of temperature on the long-range attractive interaction. Similar effects are seen for protein ions at low temperatures, although there is also some evidence for structural transitions. By heating the protein ions, their conformational profiles are significantly altered. Very high temperatures narrow the conformational space presented by both Ubiquitin and Concanavalin; it appears that diverse conformational families are "melted" into more homogeneous populations. Because of this conformational heterogeneity, the apparent IMS resolution obtained for proteins at ambient and reduced temperatures is an order of magnitude lower than the expected diffusion limited resolution (Rmax). This supports a hypothesis that the broad DTCCSHe features frequently observed for proteins do not correspond to interconverting conformers, but rather to high numbers of intrinsically stable structures.

Highlights

  • Tune Mix), the monomeric protein Ubiquitin (8.6 kDa), and the tetrameric protein complex Concanavalin A (103 kDa)

  • Instrumentation to study the fundamental properties of ion transport in gases.4. Thanks to this foundational research,5 ion mobility spectrometry (IMS) was popularized as a low cost, standalone technology for high-throughput detection of explosives and drugs.5−7 first demonstrated as an analytical separation technique by Carr in 1977,8 it was pioneered as a way to characterize isobaric species, notably by the groups of Jarrold and Bowers, in the 1990s.9−14 Data obtained from VT-Ion mobility−mass spectrometry (IM-MS) devices were instrumental in the characterization of ion−molecule interactions and subsequent developments of computational tools

  • We present our results obtained on a range of model systems, including a set of IMS standard molecules, a small protein (Ubiquitin, 8.6 kDa), and a large protein complex (Concanavalin A which presents both as a tetramer 103 kDa and a dimer 51.5 kDa)

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Summary

Analytical Chemistry

Cooling copper coil and metal encapsulated resistive heater coil, [22] LN2 port, [23] internal glass tube (shown in blue), [24] ion buncher, [25] ion buncher grids, [26] drift electrodes, [27] ceramic support ring, [28] outer ceramic rods, [29] ion funnel, [30] cone aperture, [31] hexapole lens. (C). This small offset helps to bring all the ions into the proximity of the aperture (Figure 2C(a)), where a significant hydrodynamic gas flow “blows” the ions out into the MS interface This compensates for the “trapping” and “RF heating” effects often observed in ion funnels when the bore diameter becomes close to the thickness of the electrode/spacer. IMS-ToF acquisition is versatile, as we acquire the ATD of an entire mass range It comes at a price in order to acquire high m/z data, the ToF pusher interval needs to be sufficiently long so that the high m/z ions reach the detector before the push, and when the low pressure/high field operation is desired, drift times become short and IMS peaks have widths of only a few. We measured the DTCCSHe of a 1222 m/z ion at 153 K, in two E/N regimes and using two pumping arrangements (Figure 4)

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