title: Characterization of the N-deacetylase domain from the heparan sulfate N-deacetylase/N-sulfotransferase 2.
abstract: Heparin and heparan sulfate are linear sulfated polysaccharides that exert a multitude of biological functions. Heparan sulfate glucosaminyl N-deacetylase/N-sulfotransferase isoform 2 (NDST-2), a key enzyme in the biosynthesis of heparin, contains two distinct activities. This bifunctional enzyme removes the acetyl group from N-acetylated glucosamine (N-deacetylase activity) and transfers a sulfuryl group to the unsubstituted amino position (N-sulfotransferase activity). The N-sulfotransferase activity of NDST has been unambiguously localized to the C-terminal domain of NDST. Here, we report that the N-terminal domain of NDST-2 retains N-deacetylase activity. The N-terminal domain (A66-P604) of human NDST-2, designated as N-deacetylase (NDase), was cloned as a (His)(6)-fusion protein, and protein expression was carried out in Escherichia coli. Heparosan treated with NDase contains N-unsubstituted glucosamine and is highly susceptible to N-sulfation by N-sulfotransferase. Our results conclude that the N-terminal domain of NDST-2 contains functional N-deacetylase activity. This finding helps further elucidate the mechanism of action of heparan sulfate N-deacetylase/N-sulfotransferases and the biosynthesis of heparan sulfate in general.
authors.affiliation: Division of Medicinal Chemistry and Natural Products, Rm. 309 Beard Hall, School of Pharmacy, University of North Carolina, Chapel Hill, NC 27599, USA.
authors.forename: Michael B
authors.initials: MB
authors.lastname: Duncan
authors.forename: May
authors.initials: M
authors.lastname: Liu
authors.forename: Courtney
authors.initials: C
authors.lastname: Fox
authors.forename: Jian
authors.initials: J
authors.lastname: Liu
mesh_terms: D001665:Binding Sites; D004789:Enzyme Activation; D004926:Escherichia coli / Q000201:enzymology*; D006497:Heparan Sulfate / Q000737:chemistry* / Q000378:metabolism*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D013329:Structure-Activity Relationship; D015238:Sulfotransferases / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D006497:Heparan Sulfate; D015238:Sulfotransferases; C021350:heparitin sulfotransferase