@article{butler_dodds_flick_campuzano_baker_2022, title={High-Resolution Demultiplexing (HRdm) Ion Mobility Spectrometry-Mass Spectrometry for Aspartic and Isoaspartic Acid Determination and Screening}, volume={94}, ISSN={["1520-6882"]}, DOI={10.1021/acs.analchem.1c05533}, abstractNote={Isomeric peptide analyses are an analytical challenge of great importance to therapeutic monoclonal antibody and other biotherapeutic product development workflows. Aspartic acid (Asp, D) to isoaspartic acid (isoAsp, isoD) isomerization is a critical quality attribute (CQA) that requires careful control, monitoring, and quantitation during the drug discovery and production processes. While the formation of isoAsp has been implicated in a variety of disease states such as autoimmune diseases and several types of cancer, it is also understood that the formation of isoAsp results in a structural change impacting efficacy, potency, and immunogenic properties, all of which are undesirable. Currently, lengthy ultrahigh-performance liquid chromatography (UPLC) separations are coupled with MS for CQA analyses; however, these measurements often take over an hour and drastically limit analysis throughput. In this manuscript, drift tube ion mobility spectrometry-mass spectrometry (DTIMS-MS) and both a standard and high-resolution demultiplexing approach were utilized to study eight isomeric Asp and isoAsp peptide pairs. While the limited resolving power associated with the standard DTIMS analysis only separated three of the eight pairs, the application of HRdm distinguished seven of the eight and was only unable to separate DL and isoDL. The rapid high-throughput HRdm DTIMS-MS method was also interfaced with both flow injection and an automated solid phase extraction system to present the first application of HRdm for isoAsp and Asp assessment and demonstrate screening capabilities for isomeric peptides in complex samples, resulting in a workflow highly suitable for biopharmaceutical research needs.}, number={16}, journal={ANALYTICAL CHEMISTRY}, author={Butler, Karen E. and Dodds, James N. and Flick, Tawnya and Campuzano, Iain D. G. and Baker, Erin S.}, year={2022}, month={Apr}, pages={6191–6199} }
 @article{butler_takinami_rainczuk_baker_roberts_2021, title={Utilizing Ion Mobility-Mass Spectrometry to Investigate the Unfolding Pathway of Cu/Zn Superoxide Dismutase}, volume={9}, ISSN={["2296-2646"]}, DOI={10.3389/fchem.2021.614595}, abstractNote={Native mass spectrometry has emerged as a powerful tool for structural biology as it enables the evaluation of molecules as they occur in their physiological conditions. Ion mobility spectrometry-mass spectrometry (IMS-MS) has shown essential in these analyses as it allows the measurement of the shape of a molecule, denoted as its collision cross section (CCS), and mass. The structural information garnered from native IMS-MS provides insight into the tertiary and quaternary structure of proteins and can be used to validate NMR or crystallographic X-ray structures. Additionally, due to the rapid nature (millisecond measurements) and ability of IMS-MS to analyze heterogeneous solutions, it can be used to address structural questions not possible with traditional structural approaches. Herein, we applied multiple solution conditions to systematically denature bovine Cu/Zn-superoxide dismutase (SOD1) and assess its unfolding pathway from the holo-dimer to the holo-monomer, single-metal monomer, and apo-monomer. Additionally, we compared and noted 1–2% agreement between CCS values from both drift tube IMS and trapped IMS for the SOD1 holo-monomer and holo-dimer. The observed CCS values were in excellent agreement with computational CCS values predicted from the homo-dimer crystal structure, showcasing the ability to use both IMS-MS platforms to provide valuable structural information for molecular modeling of protein interactions and structural assessments.}, journal={FRONTIERS IN CHEMISTRY}, author={Butler, Karen E. and Takinami, Yoshihiko and Rainczuk, Adam and Baker, Erin S. and Roberts, Blaine R.}, year={2021}, month={Feb} }
 @article{butler_kalmar_muddiman_baker_2021, title={Utilizing liquid chromatography, ion mobility spectrometry, and mass spectrometry to assess INLIGHT (TM) derivatized N-linked glycans in biological samples}, volume={8}, ISSN={["1618-2650"]}, DOI={10.1007/s00216-021-03570-7}, abstractNote={Glycosylation is a ubiquitous co- and post-translational modification involved in the sorting, folding, and trafficking of proteins in biological systems; in humans, >50% of gene products are glycosylated with the cellular machinery of glycosylation compromising ~2% of the genome. Perturbations in glycosylation have been implicated in a variety of diseases including neurodegenerative diseases and certain types of cancer. However, understanding the relationship between a glycan and its biological role is often difficult due to the numerous glycan isomers that exist. To address this challenge, nanoflow liquid chromatography, ion mobility spectrometry, and mass spectrometry (nLC-IMS-MS) were combined with the Individuality Normalization when Labeling with the Isotopic Glycan Hydrazide Tags (INLIGHT™) strategy to study a series of glycan standards and those enzymatically released from the glycoproteins horseradish peroxidase, fetuin, and pooled human plasma. The combination of IMS and the natural (NAT) and stable-isotope label (SIL) in the INLIGHT™ strategy provided additional confidence for each glycan identification due to the mobility aligned NAT- and SIL-labeled glycans and further capabilities for isomer examinations. Additionally, molecular trend lines based on the IMS and MS dimensions were investigated for the INLIGHT™ derivatized glycans, facilitating rapid identification of putative glycans in complex biological samples.}, journal={ANALYTICAL AND BIOANALYTICAL CHEMISTRY}, author={Butler, Karen E. and Kalmar, Jaclyn Gowen and Muddiman, David C. and Baker, Erin S.}, year={2021}, month={Aug} }
 @article{kalmar_butler_baker_muddiman_2020, title={Enhanced protocol for quantitativeN-linked glycomics analysis using Individuality Normalization when Labeling with Isotopic Glycan Hydrazide Tags (INLIGHT)(TM)}, volume={412}, ISSN={["1618-2650"]}, DOI={10.1007/s00216-020-02892-2}, abstractNote={The analysis of N-linked glycans using liquid chromatography and mass spectrometry (LC-MS) presents significant challenges, particularly owing to their hydrophilic nature. To address these difficulties, a variety of derivatization methods have been developed to facilitate improved ionization and detection sensitivity. One such method, the Individuality Normalization when Labeling with Isotopic Glycan Hydrazide Tags (INLIGHT)™ strategy for labeling glycans, has previously been utilized in the analysis of N- and O-linked glycans in biological samples. To assess the maximum sensitivity and separability of the INLIGHT™ preparation and analysis pipeline, several critical steps were investigated. First, recombinant and nonrecombinant sources of PNGase F were compared to assess variations in the released glycans. Second, modifications in the INLIGHT™ derivatization step were evaluated including temperature optimization, solvent composition changes, reaction condition length and tag concentration. Optimization of the modified method resulted in 20–100 times greater peak areas for the detected N-linked glycans in fetuin and horseradish peroxidase compared with the standard method. Furthermore, the identification of low-abundance glycans, such as (Fuc)1(Gal)2(GlcNAc)4(Man)3(NeuAc)1 and (Gal)3(GlcNAc)5(Man)3(NeuAc)3, was possible. Finally, the optimal LC setup for the INLIGHT™ derivatized N-linked glycan analyses was found to be a C18 reverse-phase (RP) column with mobile phases typical of RPLC.}, number={27}, journal={ANALYTICAL AND BIOANALYTICAL CHEMISTRY}, author={Kalmar, Jaclyn Gowen and Butler, Karen E. and Baker, Erin S. and Muddiman, David C.}, year={2020}, month={Nov}, pages={7569–7579} }