title: Enzymatic analysis of the protein O-glycosyltransferase, Rumi, acting toward epidermal growth factor-like (EGF) repeats.
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.
authors.affiliation: Department of Biochemistry and Cell Biology, Institute of Cell and Developmental Biology, Stony Brook University, Stony Brook, NY, USA.
authors.forename: Hideyuki
authors.initials: H
authors.lastname: Takeuchi
authors.forename: Robert S
authors.initials: RS
authors.lastname: Haltiwanger
mesh_terms: D020816:Amino Acid Motifs; D000818:Animals; D057075:Enzyme Assays / Q000379:methods*; D004815:Epidermal Growth Factor / Q000378:metabolism*; D005947:Glucose / Q000378:metabolism; D005964:Glucosyltransferases / Q000378:metabolism*; D006031:Glycosylation; D006801:Humans; D011994:Recombinant Proteins / Q000378:metabolism; D014994:Xylose / Q000378:metabolism
chemical_list: D011994:Recombinant Proteins; D004815:Epidermal Growth Factor; D014994:Xylose; D005964:Glucosyltransferases; C510651:POGLUT1 protein, human; D005947:Glucose