Abstract
Metabolic alterations have been identified as a frequent event in cancer. This is often associated with increased flux through glycolysis, and also a secondary pathway to glycolysis, hexosamine biosynthetic pathway (HBP). HBP provides substrate for N-linked glycosylation, which occurs in the endoplasmic reticulum and the Golgi apparatus. N-linked glycosylation supports protein folding and correct sorting of proteins to plasma membrane and secretion. This process generates complex glycoforms, which can be recognized by other proteins and glycosylation of receptor tyrosine kinases (RTK) can also regulate their plasma-membrane retention time. Of special interest for experimental biologists, plants produce proteins, termed lectins, which bind with high specificity to glyco-conjugates. For the purposes of molecular biology, plant lectins can be conjugated to different moieties, such as agarose beads, which enable precipitation of specifically glycosylated proteins. In this chapter, we describe in detail how to perform pull-down experiments with commercially available lectins to identify changes in the glycosylation of RTKs.
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Acknowledgements
HMI is funded by an EU FP7 Marie Curie Integrated Training Network, PRO-NEST and Norwegian Cancer Society by the Finnish Cultural Foundation. IGM is a visiting scientist at Cancer Research UK and an honorary senior visiting research fellow at the University of Cambridge. The Centre for Molecular Medicine Norway (NCMM) is funded by the Norwegian Research Council, University of Oslo, and the Health South East. IGM is also funded by the Norwegian Cancer Society, the Movember Foundation, and the National Institutes of Health (USA).
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Itkonen, H.M., Mills, I.G. (2015). Studying N-Linked Glycosylation of Receptor Tyrosine Kinases. In: Germano, S. (eds) Receptor Tyrosine Kinases. Methods in Molecular Biology, vol 1233. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1789-1_10
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DOI: https://doi.org/10.1007/978-1-4939-1789-1_10
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