We describe a set of simple devices for surface-induced dissociation of protein complexes on three instrument platforms. All of the devices use a novel yet simple split lens geometry that is minimally invasive (requiring a few mm along the ion path axis) and easier to operate than prior generations of devices. The split lens is designed to be small enough to replace the entrance lens of a Bruker FT-ICR collision cell, the dynamic range enhancement (DRE) lens of a Waters Q-IM-TOF, or the exit lens of a transfer multipole of a Thermo Scientific Extended Mass Range (EMR) Orbitrap. Despite the decrease in size and reduction in number of electrodes to 3 (from 10-12 in Gen 1 and ~6 in Gen 2), we show sensitivity improvement in a variety of cases across all platforms while also maintaining SID capabilities across a wide mass and energy range. The coupling of SID, high resolution, and ion mobility is demonstrated for a variety of protein complexes of varying topologies.

A second-generation (“Gen 2”) device capable of surface-induced dissociation (SID) and collision-induced dissociation (CID) for Fourier transform ion cyclotron resonance mass spectrometry of protein complexes has been designed, simulated, fabricated, and experimentally compared to a first-generation device (“Gen 1”). The primary goals of the redesign were to 1) simplify SID by reducing the number of electrodes, 2) increase CID and SID sensitivity by lengthening the collision cell, and 3) increase the mass range of the device for analysis of larger multimeric proteins, all while maintaining the normal instrument configuration and operation. Compared to Gen 1, Gen 2 exhibits an approximately 10x increase in sensitivity in flythrough mode, 7x increase in CID sensitivity for protonated leucine enkephalin (m/z 556) and 14x increase of CID sensitivity of 53 kDa streptavidin tetramer. It also approximately doubles of the useful mass range (from m/z 8,000 to m/z 15,000) using a rectilinear ion trap with a smaller inscribed radius or triples it (to m/z 22,000) using a hexapole collision cell and yields a 3–10x increase in SID sensitivity. We demonstrate the increased mass range and sensitivity on a variety of model molecules spanning nearly 3 orders of magnitude in absolute mass and present examples where the high resolution of the FT-ICR is advantageous for deconvoluting overlapping SID fragments.

<p>We describe a set of simple devices for surface-induced dissociation of protein complexes on three instrument platforms. All of the devices use a novel yet simple split lens geometry that is minimally invasive (requiring a few mm along the ion path axis) and easier to operate than prior generations of devices. The split lens is designed to be small enough to replace the entrance lens of a Bruker FT-ICR collision cell, the dynamic range enhancement (DRE) lens of a Waters Q-IM-TOF, or the exit lens of a transfer multipole of a Thermo Scientific Extended Mass Range (EMR) Orbitrap. The split lens used for SID is an order of magnitude smaller than the first-generation SID devices developed in our laboratory and approximately 5x smaller than the second-generation devices developed on the Q-IM-TOF and FT-ICR. Despite the decrease in size and reduction in number of electrodes to 3 (from 10-12 in Gen 1 and ~6 in Gen 2), we show sensitivity improvement in a variety of cases across all platforms while also maintaining SID capabilities across a wide mass and energy range. The coupling of SID, high resolution, and ion mobility is demonstrated for a variety of protein complexes of varying topologies.<b></b></p>

Native mass spectrometry (nMS), particularly in conjunction with gas-phase ion mobility spectrometry measurements, has proven useful as a structural biology tool for evaluating the stoichiometry, conformation, and topology of protein complexes. Here, we demonstrate the combination of trapped ion mobility spectrometry (TIMS) and surface-induced dissociation (SID) on a Bruker SolariX XR 15 T FT-ICR mass spectrometer for the structural analysis of protein complexes. We successfully performed SID on mobility-selected protein complexes, including the streptavidin tetramer and cholera toxin B with bound ligands. Additionally, TIMS−SID was employed on a mixture of the peptides desArg1 and desArg9 bradykinin to mobility-separate and identify the individual peptides. Importantly, results show that nativelike conformations can be maintained throughout the TIMS analysis. The TIMS−SID spectra are analogous to SID spectra acquired using quadrupole mass selection, indicating little measurable, if any, structural rearrangement during mobility selection. Mobility parking was used on the ion or mobility of interest and 50−200 SID mass spectra were averaged. High-quality TIMS−SID spectra were acquired over a period of 2−10 min, comparable to or slightly longer than SID coupled with ion mobility on various instrument platforms in our laboratory. The ultrahigh resolving power of the 15 T FT-ICR allowed for the identification and relative quantification of overlapping SID fragments with the same nominal m/z based on isotope patterns, and it shows promise as a platform to probe small mass differences, such as protein/ligand binding or post-translational modifications. These results represent the potential of TIMS−SID−MS for the analysis of both protein complexes and peptides.

No abstract