Ion Mobility-Mass Spectrometry Analysis of Large Protein-Protein Complexes: From Activated Ions to Polydisperse Assemblies

摘要

Separation by of ions by their mobility is being developed in our laboratory and others for the analysis of large protein assemblies. The overall aim of our work is to build an ion mobility-mass spectrometry approach that is capable of providing a high-confidence topology diagram for protein complexes of previously-unknown dimensions or architecture (figure 1). To this end, we have begun by studying protein assemblies with known quaternary structure in solution, but have quickly expanded to more challenging systems. Here, we present an overview of our current efforts to understand the structure and stability of large protein complexes in the absence of bulk solvent. We will emphasise recent work on activated protein complexes, where we observe dramatic structural transitions and include initial results for polydisperse assemblies. Ions were generated by nanoflow electrospray ionization from an aqueous solution buffered at pH 7. Ion mobility separation was carried out utilizing a T-wave drift cell, which replaces a standard rf ion guide collision cell within a triple quadrupole orthogonal time-of-flight mass spectrometer (Synapt, Waters, Milford, MA, USA). Ions are pulsed into the device and separated based on their ability to traverse the drift cell under the influence of a series of low-voltage "waves". Collision cross-sections were externally calibrated using protein ions of known ion mobility. Molecular modeling was performed using Maestro (Schrodinger, LLC) as well as a software package developed in-house, and collision cross sections were calculated using Mobcal (developed by Jarrold and coworkers).