Role of deacetylase activity of N-deacetylase/N-sulfotransferase 1 in forming N-sulfated domain in heparan sulfate [Enzymology]

June 24th, 2015 by Duo, W., Xu, Y., Pagadala, V., Pedersen, L. C., Liu, J.

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 compared with that made by the NDST-1 enzyme. Our data demonstrates the essential role of domain cooperation within NDST-1 in producing HS with specific domain structures.