Combining Ion Mobility and Cryogenic Spectroscopy for Structural and Analytical Studies of Biomolecular Ions

Ion mobility spectrometry (IMS) has become a valuable tool in biophysical and bioanalytical chemistry because of its ability to separate and characterize the structure of gas-phase biomolecular ions on the basis of their collisional cross section (CCS). Its importance has grown with the realization that in many cases, biomolecular ions retain important structural characteristics when produced in the gas phase by electrospray ionization (ESI). While a CCS can help distinguish between structures of radically different types, one cannot expect a single number to differentiate similar conformations of a complex molecule. Molecular spectroscopy has also played an increasingly important role for structural characterization of biomolecular ions. Spectroscopic measurements, particularly when performed at cryogenic temperatures, can be extremely sensitive to small changes in a molecule’s conformation and provide tight constraints for calculations of biomolecular structures. However, spectra of complex molecules can be heavily congested due to the presence of multiple stable conformations, each of which can have a distinct spectrum. This congestion can inhibit spectral analysis and complicate the extraction of structural information. Even when a single conformation is present, the conformational search process needed to match a measured spectrum with a computed structure can be overwhelming for peptides of more than a few amino acids, for example. We have recently combined ion mobility spectrometry and cryogenic ion spectroscopy (CIS) to characterize the structures of gas-phase biomolecular ions. In this Account, we illustrate how the coupling of IMS and CIS is by nature synergistic. On the one hand, IMS can be used as a conformational filter to reduce spectral congestion that arises from heterogeneous samples, facilitating structural analysis. On the other hand, highly resolved, cryogenic spectra can serve as a selective detector for IMS that can increase the effective resolution and hence the maximum number of distinct species that can be detected. Taken together, spectra and CCS measurements on the same system facilitates structural analysis and strengthens the conclusions that can be drawn from each type of data. After describing different approaches to combining these two techniques in such a way as to simplify the data obtained from each one separately, we present two examples that illustrate the type of insight gained from using spectra and CCS data together for characterizing gas-phase biomolecular ions. In one example, the CCS is used as a constraint for quantum chemical structure calculations of kinetically trapped species, where a lowest-energy criterion is not applicable. In a second example, we use both the CCS and a cryogenic infrared spectrum as a means to distinguish isomeric glycans.