title: Role of Deacetylase Activity of N-Deacetylase/N-Sulfotransferase 1 in Forming N-Sulfated Domain in Heparan Sulfate.
abstract: Heparan sulfate (HS) is a highly sulfated polysaccharide that plays important physiological roles. The biosynthesis of HS involves a series of enzymes, including glycosyltransferases (or HS polymerase), epimerase, and sulfotransferases. N-Deacetylase/N-Sulfotransferase isoform 1 (NDST-1) is a critical enzyme in this pathway. NDST-1, a bifunctional enzyme, displays N-deacetylase and N-sulfotransferase activities to convert an N-acetylated glucosamine residue to an N-sulfo glucosamine residue. Here, we report the cooperative effects between N-deacetylase and N-sulfotransferase activities. Using baculovirus expression in insect cells, we obtained three recombinant proteins: full-length NDST-1 and the individual N-deacetylase and N-sulfotransferase domains. Structurally defined oligosaccharide substrates were synthesized to test the substrate specificities of the enzymes. We discovered that N-deacetylation is the limiting step and that interplay between the N-sulfotransferase and N-deacetylase accelerates the reaction. Furthermore, combining the individually expressed N-deacetylase and N-sulfotransferase domains produced different sulfation patterns when compared with that made by the NDST-1 enzyme. Our data demonstrate the essential role of domain cooperation within NDST-1 in producing HS with specific domain structures.
authors.affiliation: From the Division of Chemical Biology and Medicinal Chemistry, Eshelman School of Pharmacy, University of North Carolina, Chapel Hill, North Carolina 27599, the Laboratory of Pharmaceutical Engineering, School of Pharmaceutical Sciences, Jiangnan University, Wuxi 214122, China, and.
authors.forename: Wenfang
authors.initials: W
authors.lastname: Dou
authors.affiliation: From the Division of Chemical Biology and Medicinal Chemistry, Eshelman School of Pharmacy, University of North Carolina, Chapel Hill, North Carolina 27599.
authors.forename: Yongmei
authors.initials: Y
authors.lastname: Xu
authors.affiliation: From the Division of Chemical Biology and Medicinal Chemistry, Eshelman School of Pharmacy, University of North Carolina, Chapel Hill, North Carolina 27599.
authors.forename: Vijayakanth
authors.initials: V
authors.lastname: Pagadala
authors.affiliation: the Genome Integrity and Structural Biology Laboratory, NIEHS, National Institutes of Health, Research Triangle Park, North Carolina 27709.
authors.forename: Lars C
authors.initials: LC
authors.lastname: Pedersen
authors.affiliation: From the Division of Chemical Biology and Medicinal Chemistry, Eshelman School of Pharmacy, University of North Carolina, Chapel Hill, North Carolina 27599, jian_liu@unc.edu.
authors.forename: Jian
authors.initials: J
authors.lastname: Liu
mesh_terms: D002240:Carbohydrate Sequence; D006497:Heparan Sulfate / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D013379:Substrate Specificity; D015238:Sulfotransferases / Q000737:chemistry / Q000378:metabolism*; D053719:Tandem Mass Spectrometry
chemical_list: D006497:Heparan Sulfate; D015238:Sulfotransferases; C021350:heparitin sulfotransferase
keywords: carbohydrate biosynthesis; heparan sulfate; heparin; oligosaccharide; sulfotransferase