Multidimensional ion mobility spectrometry techniques (IMS-IMS and IMS-IMS-IMS) combined with mass spectrometry are used to study structural transitions of ubiquitin ions in the gas phase. It is possible to select and activate narrow distributions of compact and partially folded conformation types and examine new distributions of structures that are formed. Different compact conformations unfold, producing a range of new partially folded states and three resolvable peaks associated with elongated conformers. Under gentle activation conditions, the final populations of the three elongated forms depend on the initial structures of the selected ions. This requires that some memory of the compact state (most likely secondary structure) is preserved along the unfolding pathway. Activation of selected, partially folded intermediates (formed from specific compact states) leads to elongated state populations that are consistent with the initial selected compact form-evidence that intermediates not only retain elements of initial structure but also are capable of transmitting structure to final states.

In this report, we describe a dual ionization source ion mobility-mass spectrometer (IM-MS) instrument platform for investigations that critically compare ion mobility collision cross section (CCS) measurements obtained from different ionization methods. The instrument incorporates both matrix-assisted laser desorption ionization (MALDI) and nano-electrospray ionization (nESI) sources. The nESI source incorporates a keyhole geometry ion funnel design which facilitates axial ion focusing, accumulation, and generation of short duration (10–30 µs) ion pulses for use with the IM-MS. The IM-MS instrument operation is independent of which ionization source is used. This allows comparisons of collision cross section measurements to be made between both ion sources with minimal differences in the instrumental arrangement. The performance of the nESI ion source is evaluated by measuring the collision cross section values of the charge states of equine cytochrome c (z = 9 to 16) and values are in good agreement (<2% deviation) with those previously reported in the literature. Several charge states (z = 8 to 11) of cytochrome c exhibit multiple cross sectional features in the ion mobility analysis. An analysis of the tryptic peptides of cytochrome c formed by both ESI and MALDI demonstrate that on average, +1 MALDI ions are similar in CCS to +1 ESI ions and are smaller than +2 ESI ions. The ion mobility resolving power with ESI (30–35) is comparable to that obtained by using MALDI (35–40), which suggests that both sources produce sufficiently narrow ion pulses for the measurement to be predominately diffusion rather than gate pulse width limited.

No abstract