Heparan/Chondroitin Sulfate Biosynthesis

Pedersen, Lars C.; Tsuchida, Kazunori; Kitagawa, Hiroshi; Sugahara, Kazuyuki; Darden, Thomas A.; Negishi, Masahiko

doi:10.1074/jbc.m007399200

Cited by 175 publications

(84 citation statements)

References 29 publications

(17 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In fact, we have observed that the synthetic phosphorylated trisaccharideserine, Gal-Gal-Xyl(2-O-phosphate)-Ser, served as a better acceptor substrate than the unmodified counterpart for recombinant human GlcUA transferase I. 2 The acceptor recognition by a crystallized form of this GlcUA transferase I has recently been demonstrated by x-ray crystallography (55), and it has further been revealed that the crystallized enzyme specifically recognized the 6-O-sulfate group on the Gal(I) residue adjacent to the Xyl residue, 2 although it remains to be determined whether the phosphate on the Xyl and/or the 4-O-sulfate group on the Gal(II) residue can also be recognized by the crystallized enzyme. These findings suggest that the phosphorylation of the Xyl residue and sulfation of the Gal residues may be required for efficient elongation and maturation of the linkage region tetrasaccharide as prerequisites for the assembly of GAG chains.…”

Section: Discussionmentioning

confidence: 99%

Structural Characterization of Heparan Sulfate and Chondroitin Sulfate of Syndecan-1 Purified from Normal Murine Mammary Gland Epithelial Cells

Ueno¹,

Yamada²,

Zako³

et al. 2001

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

Syndecan-1, present on the surfaces of normal murine mammary gland epithelial cells, is a transmembrane hybrid proteoglycan, which bears glycosaminoglycan (GAG) side chains of heparan sulfate (HS) and chondroitin sulfate (CS). Purified syndecan-1 ectodomains were analyzed for disaccharide composition and the GAG-protein linkage region after digestion with bacterial lyases. The HS chains contained predominantly a nonsulfated unit with smaller proportions of two monosulfated, two disulfated, and a trisulfated unit, whereas CS chains were demonstrated for the first time to bear GlcUA-GalNAc(4-O-sulfate) as a major component as well as GlcUA-GalNAc, GlcUA-GalNAc(6-O-sulfate), and an E disaccharide unit GlcUA-GalNAc(4,6-O-disulfate) as minor yet appreciable components. Two kinds of linkage region tetrasaccharides, GlcUA-Gal-Gal-Xyl and GlcUA-Gal-Gal-Xyl(2-O-phosphate), were found for the HS chains in a molar ratio of 55:45. In marked contrast, an additional sulfated tetrasaccharide, GlcUA-Gal(4-Osulfate)-Gal-Xyl, was demonstrated only for the CS chains, and the unmodified phosphorylated and sulfated components were present at a molar ratio of 55:26: 19. The present study thus provided conclusive evidence for the hypothesis that 4-O-sulfation of Gal is peculiar to CS chains in contrast to the phosphorylation of Xyl, which is common to both HS and CS chains. These modifications may be required for biosynthetic maturation of the linkage region tetrasaccharide sequence, which is a prerequisite for creating the repeating disaccharide region of GAG chains and/or biosynthetic selective chain assembly of CS and HS chains.

show abstract

Section: Discussionmentioning

confidence: 99%

Structural Characterization of Heparan Sulfate and Chondroitin Sulfate of Syndecan-1 Purified from Normal Murine Mammary Gland Epithelial Cells

Ueno¹,

Yamada²,

Zako³

et al. 2001

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

show abstract

“…In addition, DVD (motif ii) participates in coordination of Mn ϩϩ , the small amino acids (GG) within motif iii allow the donor sugar to fit deep into the pocket, and an acidic amino acid within motif iv is proposed to make contact with the sugar acceptor and to participate in nucleophilic attack on it. Similarly, in the crystallographic structures of human GlcAT-I with its substrates (26,27), DDD (motif ii) interacts with UDP and Mn ϩϩ , and the E within motif iv interacts with the acceptor substrate and is proposed to participate directly in catalysis. Indeed, even though bovine ␤4GalT1 and human GlcAT-I exhibit no significant primary sequence similarity, when the locations of conserved motifs ii, iii, and iv are mapped onto their crystallographic structures, it is evident that the spatial relationships of these motifs to each other and to the substrate binding pocket are similar (Fig.…”

Section: Resultsmentioning

confidence: 99%

Molecular genetic analysis of the glycosyltransferase Fringe in Drosophila

Correia

Papayannopoulos

Panin

et al. 2003

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

Fringe proteins are ␤1,3-N-acetylglucosaminyltransferases that modulate signaling through Notch receptors by modifying Olinked fucose on epidermal growth factor domains. Fringe is highly conserved, and comparison among 18 different Fringe proteins from 11 different species identifies a core set of 84 amino acids that are identical among all Fringes. Fringe is only distantly related to other glycosyltransferases, but analysis of the predicted Drosophila proteome identifies a set of four sequence motifs shared among Fringe and other putative ␤1,3-glycosyltransferases. To gain functional insight into these conserved sequences, we genetically and molecularly characterized 14 point mutations in Drosophila fringe. Most nonsense mutations act as recessive antimorphs, raising the possibility that Fringe may function as a dimer. Missense mutations identify two distinct motifs that are conserved among ␤1,3-glycosyltransferases, and that can be modeled onto key motifs in the crystallographic structures of bovine ␤1,4-galactosyltransferase 1 and human glucuronyltransferase I. Other missense mutations map to amino acids that are conserved among Fringe proteins, but not among other glycosyltransferases, and thus may identify structural motifs that are required for unique aspects of Fringe activity. Although the Notch pathway mediates a wide range of cell fate decisions, the influence of fringe ( fng) genes on Notch signaling has been best studied during the development of the Drosophila wing. In the developing wing imaginal disk, FNG inhibits the ability of Serrate to activate Notch, and potentiates the ability of Delta to activate Notch. These effects of FNG position a stripe of Notch activation along the border between dorsal and ventral cells, which is essential for the subsequent patterning, growth, and compartmentalization of the wing (reviewed in refs. 5 and 6).Drosophila and vertebrate FNG proteins are highly conserved and possess an unusual ␤1,3-N-acetylglucosaminyltransferase activity that elongates O-linked fucose within EGF domains (7,8). Although FNG is not closely related to other glycosyltransferases, it does share a few short sequence motifs with certain other glycosyltransferases (9), and we report here that it is most closely related to members of a ␤1,3-glycosyltransferase (␤3GT) superfamily. These enzymes use a variety of sugar donors and acceptors, but all transfer the sugar from a UDP-sugar donor, creating a ␤ linkage between the 1 carbon of the donor and the 3 carbon of the acceptor. With the exception of site-directed mutations in a DDD motif of FNG (7,8,10,38), the functional requirements for conserved sequence motifs in FNG or other ␤3GTs have not been assessed. Moreover, the only crystallographic structure for a ␤3GT determined to date is that of human glucuronyltransferase I (GlcAT-I), which does not share significant primary sequence similarity with FNG.Here, we employ two complementary approaches to identify functionally important amino acids within FNG: sequence conservation, and genetic and mole...

show abstract

“…: 919-541-2404; Fax: 919-541-0696; E-mail: negishi@niehs.nih.gov. 1 The abbreviations used are: GlcAT-I, ␤1,3-glucuronyltransferase; NDST, N-deacetylase/N-sulfotransferase; MES, 4-morpholineethanesulfonic acid; MME-PEG-2000, monomethyl ether polyethylene glycol 2000. 100 mM MES, pH 6.0, 10 mM MnCl 2 , 10 mM UDP-GlcUA, and 10% ethylene glycol.…”

Section: Methodsmentioning

confidence: 99%

“…Protein of the recombinant human GlcAT-I catalytic domain was cloned, expressed, and purified as previously described (1). Crystals of the enzyme were obtained by the vapor diffusion hanging drop method.…”

Section: Methodsmentioning

confidence: 99%