The Development and Utilization of the Periodic Focusing Ion Funnel

Ion mobility-mass spectrometry (IM-MS) provides gas phase, size-based separation, on an ultrafast timescale (?s-ms). With the incorporation of electrospray ionization, IM-MS is a valuable tool to investigate conformations of biological molecule ions that can be representative of their solution-phase structure. In some cases, evaporative cooling during ESI can kinetically trap these solution-phase structures in local minima along the potential energy surface. However, if the internal energy of the ion is increased via collisional activation, these solution-phase structures can be readily converted to an energetically preferred, gas-phase structure. Radio frequency (RF) confining devices, such as the RF ion funnel, are typically used to increase ion transmission in IM-MS measurements; however, these devices can lead to collisional activation and structural rearrangement due to high voltage oscillation amplitudes (Vp-p). Recently, periodic focusing ion mobility spectrometry (PF IMS) has been shown to provide comparable radial confinement, while utilizing reduced radial electric fields Vp-p as compared to the RF ion funnel. Work presented herein describes the development and characterization of a periodic focusing ion funnel (PF IF) that is capable of increasing ion transmission while being able to preserve nascent conformer distributions and subsequently inducing structural rearrangement.

The utility of the PF IF is demonstrated with the neuropeptide Substance P (SP), as it provides a model for studying the structural effects of collisional activation due to the presence of both a kinetically trapped and gas-phase conformer, denoted ASP and BSP, respectively. By increasing the internal energy of [SP + 3H]^3+ ions, ASP is quantitatively converted to BSP, which is consistent with ASP being a kinetically trapped conformer and BSP being a gas-phase conformer. The collision cross section and mobility resolution of the ASP suggests that it is comprised of a broad distribution of compact globular conformations. Intramolecular solvation appears to stabilize the compacted structure of ASP in the gas-phase; however, as the ion?s internal energy increases, these noncovalent interactions are disrupted and the peptide converts into the gas-phase conformation. Mutations of various amino acid residues of SP provide a means of identifying these interactions and their effect on the stability of the kinetically trapped conformers.