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Tandem-trapped ion mobility spectrometry/mass spectrometry (tTIMS/MS): a promising analytical method for investigating heterogenous samples.

ABSTRACT: Ion mobility spectrometry/mass spectrometry (IMS/MS) is widely used to study various levels of protein structure. Here, we review the current state of affairs in tandem-trapped ion mobility spectrometry/mass spectrometry (tTIMS/MS). Two different tTIMS/MS instruments are discussed in detail: the first tTIMS/MS instrument, constructed from coaxially aligning two TIMS devices; and an orthogonal tTIMS/MS configuration that comprises an ion trap for irradiation of ions with UV photons. We discuss the various workflows the two tTIMS/MS setups offer and how these can be used to study primary, tertiary, and quaternary structures of protein systems. We also discuss, from a more fundamental perspective, the processes that lead to denaturation of protein systems in tTIMS/MS and how to soften the measurement so that biologically meaningful structures can be characterised with tTIMS/MS. We emphasize the concepts underlying tTIMS/MS to underscore the opportunities tandem-ion mobility spectrometry methods offer for investigating heterogeneous samples.

SUBMITTER: Liu FC

PROVIDER: S-EPMC9914546 | biostudies-literature | 2022 May

REPOSITORIES: biostudies-literature

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Tandem-trapped ion mobility spectrometry/mass spectrometry (tTIMS/MS): a promising analytical method for investigating heterogenous samples.

Liu Fanny C FC Ridgeway Mark E ME Park Melvin A MA Bleiholder Christian C

The Analyst 20220530 11

Ion mobility spectrometry/mass spectrometry (IMS/MS) is widely used to study various levels of protein structure. Here, we review the current state of affairs in tandem-trapped ion mobility spectrometry/mass spectrometry (tTIMS/MS). Two different tTIMS/MS instruments are discussed in detail: the first tTIMS/MS instrument, constructed from coaxially aligning two TIMS devices; and an orthogonal tTIMS/MS configuration that comprises an ion trap for irradiation of ...[more]

PMID: 35521797

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Project description:Ion mobility-mass spectrometry (IM-MS) has become a powerful tool for glycan structural characterization due to its ability to separate isomers and provide collision cross section (CCS) values that facilitate structural assignment. However, IM-based isomer analysis may be complicated by the presence of multiple gas-phase conformations of a single structure that not only increases difficulty in isomer separation but can also introduce the possibility for misinterpretation of conformers as isomers. Here, the ion mobility behavior of several sets of isomeric glycans, analyzed as their permethylated derivatives, in both nonreduced and reduced forms, was investigated by gated-trapped ion mobility spectrometry (G-TIMS). Notably, reducing-end reduction, commonly performed to remove anomerism-induced chromatographic peak splitting, did not eliminate the conformational heterogeneity of permethylated glycans in the gas phase. At a mobility resolving power of ∼100, 14 out of 22 structures showed more than one conformation. These results highlight the need to use IMS devices with high mobility resolving power for better separation of isomers and to acquire additional structural information that can differentiate isomers from conformers. Online electronic excitation dissociation (EED) MS/MS analysis of isomeric glycan mixtures following G-TIMS separation showed that EED can generate isomer-specific fragments while producing nearly identical tandem mass spectra for conformers, thus allowing confident identification of isomers with minimal evidence of any ambiguity resulting from the presence of conformers. G-TIMS EED MS/MS analysis of N-linked glycans released from ovalbumin revealed that several mobility features previously thought to arise from isomeric structures were conformers of a single structure. Finally, analysis of ovalbumin N-glycans from different sources showed that the G-TIMS EED MS/MS approach can accurately determine the batch-to-batch variations in glycosylation profiles at the isomer level, with confident assignment of each isomeric structure.

Dataset Information

Tandem-trapped ion mobility spectrometry/mass spectrometry (tTIMS/MS): a promising analytical method for investigating heterogenous samples.

Publications

Tandem-trapped ion mobility spectrometry/mass spectrometry (<i>t</i>TIMS/MS): a promising analytical method for investigating heterogenous samples.

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