Biophys J, December 2000, p. 2909-2917, Vol. 79, No. 6

Heparan Sulfate Biosynthesis: A Theoretical Study of the Initial Sulfation Step by N-Deacetylase/N-Sulfotransferase

Anna Gorokhov,^* Lalith Perera, Thomas A. Darden,^* Masahiko Negishi,^* Lars C. Pedersen,^* and Lee G. Pedersen^*

 ^*National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina 27709 and  Department of Chemistry, University of North Carolina, Chapel Hill, North Carolina 27599-3290 USA

Heparan sulfate N-deacetylase/N-sulfotransferase (NDST) catalyzes the deacetylation and sulfation of N-acetyl-D-glucosamine residues of heparan sulfate, a key step in its biosynthesis. Recent crystallographic and mutational studies have identified several potentially catalytic residues of the sulfotransferase domain of this enzyme (Kakuta et al., 1999, J. Biol. Chem. 274:10673-10676). We have used the x-ray crystal structure of heparan sulfate N-sulfotransferase with 3'-phosphoadenosine 5'-phosphate to build a solution model with cofactor 3'-phosphoadenosine 5'-phosphosulfate (PAPS) and a model heparan sulfate ligand bound, and subsequently performed a 2-ns dynamics solution simulation. The simulation results confirm the importance of residues Glu⁶⁴², Lys⁶¹⁴, and Lys⁸³³, with the possible involvement of Thr⁶¹⁷ and Thr⁶¹⁸, in binding PAPS. Additionally, Lys⁶⁷⁶ is found in close proximity to the reaction site in our solvated structure. This study illustrates for the first time the possible involvement of water in the catalysis. Three water molecules were found in the binding site, where they are coordinated to PAPS, heparan sulfate, and the catalytic residues.

Biophys J, December 2000, p. 2909-2917, Vol. 79, No. 6
© 2000 by the Biophysical Society   0006-3495/00/12/2909/09  $2.00

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