Glycoform analysis of intact erythropoietin by MALDI FT-ICR mass spectrometry
| Autor: | Steffen Lippold, Raashina Thavarajah, Dietmar Reusch, Simone Nicolardi, Manfred Wuhrer |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2021 |
| Předmět: |
Glycan
Spectrometry Mass Electrospray Ionization Glycosylation 010504 meteorology & atmospheric sciences Resolution (mass spectrometry) Mass spectrometry 01 natural sciences Biochemistry Analytical Chemistry law.invention Biopharmaceuticals chemistry.chemical_compound MALDI FT-ICR MS law hemic and lymphatic diseases medicine Environmental Chemistry Humans Erythropoietin Spectroscopy 0105 earth and related environmental sciences chemistry.chemical_classification biology 010401 analytical chemistry Intact protein Recombinant Proteins Sialic acid 0104 chemical sciences carbohydrates (lipids) chemistry Spectrometry Mass Matrix-Assisted Laser Desorption-Ionization Recombinant DNA biology.protein Glycoprotein medicine.drug |
| Zdroj: | Analytica Chimica Acta, 1185. ELSEVIER Analytica chimica acta |
| Popis: | Recombinant human erythropoietin (EPO) is a complex therapeutic glycoprotein with three N- and one O-glycosylation sites. Glycosylation of EPO influences its safety and efficacy and is defined as a critical quality attribute. Thus, analytical methods for profiling EPO glycosylation are highly demanded. Owing to the complexity of the intact protein, information about EPO glycosylation is commonly derived from released glycan and glycopeptide analysis using mass spectrometry (MS). Alternatively, comprehensive insights into the glycoform heterogeneity of intact EPO are obtained using ESI MS-based methods with or without upfront separation of EPO glycoforms. MALDI MS, typically performed with TOF mass analyzers, has been also used for the analysis of intact EPO but, due to the poor glycoform resolution, has only provided limited glycoform information. Here, we present a MALDI FT-ICR MS method for the glycosylation profiling of intact EPO with improved glycoform resolution and without loss of sialic acid residues commonly observed in MALDI analysis. Three EPO variants were characterized in-depth and up to 199 glycoform compositions were assigned from the evaluation of doubly-charged ions, without any deconvolution of the mass spectra. Key glycosylation features such as sialylation, acetylation, and N- acetyllactosamine repeats were determined and found to agree with previously reported data obtained from orthogonal analyses. The developed method allowed for a fast and straightforward data acquisition and evaluation and can be potentially used for the high-throughput comparison of EPO samples throughout its manufacturing process. (c) 2021 The Author(s). Published by Elsevier B.V. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/). |
| Databáze: | OpenAIRE |
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