Native and Denaturing MS Protein Deconvolution for Biopharma: Monoclonal Antibodies and Antibody–Drug Conjugates to Polydisperse Membrane Proteins and Beyond
| Autor: | John O. Hui, Stone D.-H. Shi, Pascal F. Egea, John H. Robinson, Jennifer L. Lippens, Michael Nshanian, Dhanashri Bagal, Joseph A. Loo, Marshall Bern, Chawita Netirojjanakul, Iain D. G. Campuzano |
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| Rok vydání: | 2019 |
| Předmět: |
Spectrometry
Mass Electrospray Ionization Immunoconjugates Glycosylation Electrospray ionization Bioengineering 010402 general chemistry Mass spectrometry Orbitrap 01 natural sciences Article Antibodies Biopharmaceutics Analytical Chemistry law.invention law Monoclonal Nanodisc Chromatography Spectrometry Chemistry Lysine Electrospray Ionization 010401 analytical chemistry Antibodies Monoclonal Proteins Membrane Proteins Mass 0104 chemical sciences Molecular Weight Membrane protein Membrane protein complex Deconvolution Other Chemical Sciences Algorithms Ion cyclotron resonance Biotechnology |
| Zdroj: | Analytical chemistry, vol 91, iss 15 Anal Chem |
| ISSN: | 1520-6882 0003-2700 |
| DOI: | 10.1021/acs.analchem.9b00062 |
| Popis: | Electrospray ionization mass spectrometry (ESI-MS) is a ubiquitously used analytical method applied across multiple departments in biopharma, ranging from early research discovery to process development. Accurate, efficient, and consistent protein MS spectral deconvolution across multiple instrument and detector platforms (time-of-flight, Orbitrap, Fourier-transform ion cyclotron resonance) is essential. When proteins are ionized during the ESI process, a distribution of consecutive multiply charged ions are observed on the m/z scale, either positive [M + nH]n+ or negative [M - nH]n- depending on the ionization polarity. The manual calculation of the neutral molecular weight (MW) of single proteins measured by ESI-MS is simple; however, algorithmic deconvolution is required for more complex protein mixtures to derive accurate MWs. Multiple deconvolution algorithms have evolved over the past two decades, all of which have their advantages and disadvantages, in terms of speed, user-input parameters (or ideally lack thereof), and whether they perform optimally on proteins analyzed under denatured or native-MS and solution conditions. Herein, we describe the utility of a parsimonious deconvolution algorithm (explaining the observed spectra with a minimum number of masses) to process a wide range of highly diverse biopharma relevant and research grade proteins and complexes (PEG-GCSF; an IgG1k; IgG1- and IgG2-biotin covalent conjugates; the membrane protein complex AqpZ; a highly polydisperse empty MSP1D1 nanodisc and the tetradecameric chaperone protein complex GroEL) analyzed under native-MS, denaturing LC-MS, and positive and negative modes of ionization, using multiple instruments and therefore multiple data formats. The implementation of a comb filter and peak sharpening option is also demonstrated to be highly effective for deconvolution of highly polydisperse and enhanced separation of a low level lysine glycation post-translational modification (+162.1 Da), partially processed heavy chain lysine residues (+128.1 Da), and loss of N-acetylglucosamine (GlcNAc; -203.1 Da). |
| Databáze: | OpenAIRE |
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