Glycobiology, 2002, Vol. 12, No. 11 763-770
© 2002 Oxford University Press
O-Glycosylation of EGF repeats: identification and initial characterization of a UDP-glucose: protein O-glucosyltransferase
3 Department of Biochemistry and Cell Biology, Institute for Cell and Developmental Biology, State University of New York at Stony Brook, Stony Brook, NY 11794-5215, USA
O-Glucose is an unusual form of posttranslational modification consisting of glucose directly attached to protein through O-linkage. Several serum proteins (factor VII, factor IX, protein Z, and thrombospondin) contain this unique modification on their epidermal growth factor (EGF)–like repeats. Comparison of the glycosylation sites on these proteins revealed a putative consensus sequence for O-glucose modification: C1XSXPC2, where C1 and C2 are the first and second conserved cysteines of the EGF repeat. We identify and characterize an enzymatic activity capable of adding glucose to EGF repeats: UDP-glucose: protein O-glucosyltransferase. Using extracts of Chinese hamster ovary cells as the enzyme source, recombinant factor VII EGF repeat as the acceptor, and UDP-[3H]glucose as the donor, we show that the activity is linearly dependent on time, enzyme amount, and substrate concentration. As with most glycosyltransferases, metal ions (such as manganese) are required for activity. Analysis demonstrated that the glucose is added in O-linkage to the EGF repeat. Mutation of the serine to alanine in the predicted glycosylation site abrogates glycosylation, as does reduction and alkylation of the EGF repeat, suggesting that the enzyme recognizes not only the consensus sequence but also the 3D structure of the EGF repeat. Detection of O-glucosyltransferase activity in extracts of cell lines from insects to humans and a variety of rat tissues suggests that O-glucose modification is widespread in biology. These studies lay the foundation for future work on the biological role of the O-glucose modification.
1 Present address: Department of Cell Biology, Albert Einstein College of Medicine, 1300 Morris Park Ave., Bronx, NY 10461, USA
2 To whom correspondence should be addressed; E-mail: Robert.Haltiwanger@stonybrook.edu
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  T. Okajima, A. Matsuura, and T. Matsuda Biological Functions of Glycosyltransferase Genes Involved in O-fucose Glycan Synthesis J. Biochem., July 1, 2008; 144(1): 1 - 6. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. M. Gebauer, S. Muller, F.-G. Hanisch, M. Paulsson, and R. Wagener O-Glucosylation and O-Fucosylation Occur Together in Close Proximity on the First Epidermal Growth Factor Repeat of AMACO (VWA2 Protein) J. Biol. Chem., June 27, 2008; 283(26): 17846 - 17854. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. Ishimizu, K. Sano, T. Uchida, H. Teshima, K. Omichi, H. Hojo, Y. Nakahara, and S. Hase Purification and Substrate Specificity of UDP-D-xylose:{beta}-D-Glucoside {alpha}-1,3-D-Xylosyltransferase Involved in the Biosynthesis of the Xyl{alpha}1-3Xyl{alpha}1-3Glc{beta}1-O-Ser on Epidermal Growth Factor-like Domains J. Biochem., April 1, 2007; 141(4): 593 - 600. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Kozma, J. J. Keusch, B. Hegemann, K. B. Luther, D. Klein, D. Hess, R. S. Haltiwanger, and J. Hofsteenge Identification and Characterization of abeta1,3-Glucosyltransferase That Synthesizes the Glc-beta1,3-Fuc Disaccharide on Thrombospondin Type 1 Repeats J. Biol. Chem., December 1, 2006; 281(48): 36742 - 36751. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Luo, A. Nita-Lazar, and R. S. Haltiwanger Two Distinct Pathways for O-Fucosylation of Epidermal Growth Factor-like or Thrombospondin Type 1 Repeats J. Biol. Chem., April 7, 2006; 281(14): 9385 - 9392. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Luo, K. Koles, W. Vorndam, R. S. Haltiwanger, and V. M. Panin Protein O-Fucosyltransferase 2 Adds O-Fucose to Thrombospondin Type 1 Repeats J. Biol. Chem., April 7, 2006; 281(14): 9393 - 9399. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. Wopereis, D. J. Lefeber, E. Morava, and R. A. Wevers Mechanisms in Protein O-Glycan Biosynthesis and Clinical and Molecular Aspects of Protein O-Glycan Biosynthesis Defects: A Review Clin. Chem., April 1, 2006; 52(4): 574 - 600. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. Rampal, A. S. Y. Li, D. J. Moloney, S. A. Georgiou, K. B. Luther, A. Nita-Lazar, and R. S. Haltiwanger Lunatic Fringe, Manic Fringe, and Radical Fringe Recognize Similar Specificity Determinants in O-Fucosylated Epidermal Growth Factor-like Repeats J. Biol. Chem., December 23, 2005; 280(51): 42454 - 42463. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 T. Okajima, A. Xu, L. Lei, and K. D. Irvine Chaperone Activity of Protein O-Fucosyltransferase 1 Promotes Notch Receptor Folding Science, March 11, 2005; 307(5715): 1599 - 1603. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. Shao, D. J. Moloney, and R. Haltiwanger Fringe Modifies O-Fucose on Mouse Notch1 at Epidermal Growth Factor-like Repeats within the Ligand-binding Site and the Abruptex Region J. Biol. Chem., February 28, 2003; 278(10): 7775 - 7782. [Abstract] [Full Text] [PDF]
|
|