title: Crystal structure of N-glycosylated human glypican-1 core protein: structure of two loops evolutionarily conserved in vertebrate glypican-1.
abstract: Glypicans are a family of cell-surface proteoglycans that regulate Wnt, hedgehog, bone morphogenetic protein, and fibroblast growth factor signaling. Loss-of-function mutations in glypican core proteins and in glycosaminoglycan-synthesizing enzymes have revealed that glypican core proteins and their glycosaminoglycan chains are important in shaping animal development. Glypican core proteins consist of a stable α-helical domain containing 14 conserved Cys residues followed by a glycosaminoglycan attachment domain that becomes exclusively substituted with heparan sulfate (HS) and presumably adopts a random coil conformation. Removal of the α-helical domain results in almost exclusive addition of the glycosaminoglycan chondroitin sulfate, suggesting that factors in the α-helical domain promote assembly of HS. Glypican-1 is involved in brain development and is one of six members of the vertebrate family of glypicans. We expressed and crystallized N-glycosylated human glypican-1 lacking HS and N-glycosylated glypican-1 lacking the HS attachment domain. The crystal structure of glypican-1 was solved using crystals of selenomethionine-labeled glypican-1 core protein lacking the HS domain. No additional electron density was observed for crystals of glypican-1 containing the HS attachment domain, and CD spectra of the two protein species were highly similar. The crystal structure of N-glycosylated human glypican-1 core protein at 2.5 Å, the first crystal structure of a vertebrate glypican, reveals the complete disulfide bond arrangement of the conserved Cys residues, and it also extends the structural knowledge of glypicans for one α-helix and two long loops. Importantly, the loops are evolutionarily conserved in vertebrate glypican-1, and one of them is involved in glycosaminoglycan class determination.
authors.affiliation: Department of Experimental Medical Science, Division of Neuroscience, Glycobiology Group, Lund University, Biomedical Center A13, SE-221 84 Lund, Sweden.
authors.forename: Gabriel
authors.initials: G
authors.lastname: Svensson
authors.forename: Wael
authors.initials: W
authors.lastname: Awad
authors.forename: Maria
authors.initials: M
authors.lastname: Håkansson
authors.forename: Katrin
authors.initials: K
authors.lastname: Mani
authors.forename: Derek T
authors.initials: DT
authors.lastname: Logan
mesh_terms: D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D002942:Circular Dichroism; D003001:Cloning, Molecular; D017124:Conserved Sequence; D018360:Crystallography, X-Ray / Q000379:methods*; D004220:Disulfides / Q000737:chemistry; D006023:Glycoproteins / Q000737:chemistry; D006031:Glycosylation; D053673:Glypicans / Q000737:chemistry*; D057809:HEK293 Cells; D006497:Heparan Sulfate / Q000737:chemistry; D006801:Humans; D008958:Models, Molecular; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011509:Proteoglycans / Q000737:chemistry
chemical_list: D004220:Disulfides; D006023:Glycoproteins; D053673:Glypicans; D006497:Heparan Sulfate; D011509:Proteoglycans