Abstract
Glycoprofiling recombinant proteins expressed and secreted from mammalian cells is key to understanding their interactions with glycoprotein receptors in vivo. Recently, recombinant T cell receptors (TCRs) are being considered as therapeutic moieties. Here we present a mass spectrometry based protocol with a “bottom up” approach to characterize glycosylation in recombinant fusion proteins with α/β TCR constant domains expressed in mammalian cells. The protocol focuses on using peptide mass mapping and mass spectrometry for N-linked glycan profiling, including analyses of site occupancy, glycan heterogeneity, and possible glycan compositions and structures.
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Acknowledgments
The authors thank Bryan E. Jones and Wolfgang Glaesner for their managerial support, Flora Huang for the purification of TCR proteins, and Jayd Hanna and Benjamin Gutierrez for assistance with transient transfection and 293F cell culture.
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Zhang, K., Demarest, S.J., Wu, X., Fitchett, J.R. (2017). Glycosylation Profiling of α/β T Cell Receptor Constant Domains Expressed in Mammalian Cells. In: Tiller, T. (eds) Synthetic Antibodies. Methods in Molecular Biology, vol 1575. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6857-2_12
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DOI: https://doi.org/10.1007/978-1-4939-6857-2_12
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