Abstract
Epidermal growth factor-like (EGF) repeats are found in numerous extracellular or transmembrane proteins including Notch. EGF repeats containing the appropriate consensus sequences can be modified with two unusual types of glycans: O-fucosylation and O-glucosylation. We have identified the glycosyltransferases that catalyze the addition of the first sugar to these consensus sites: protein O-fucosyltransferase 1 (Pofut1) and protein O-glucosyltransferase (Rumi/Poglut1). Recently, we have demonstrated that Rumi/Poglut1 shows protein O-xylosyltransferase activity as well. Here, we describe how we characterize the enzymatic activity of these enzymes, including preparation of the acceptor substrates, using bacterially expressed EGF repeats.
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Acknowledgments
We would like to thank Haltiwanger lab members for helpful comments. The primary work introduced here was supported by NIH grant GM061126 (to RSH) and the research grant from Mizutani Foundation for Glycoscience (to HT).
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Takeuchi, H., Haltiwanger, R.S. (2013). Enzymatic Analysis of the Protein O-Glycosyltransferase, Rumi, Acting Toward Epidermal Growth Factor-Like (EGF) Repeats. In: Brockhausen, I. (eds) Glycosyltransferases. Methods in Molecular Biology, vol 1022. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-465-4_10
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DOI: https://doi.org/10.1007/978-1-62703-465-4_10
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