| Abstract
| - The 3-O-sulfation of glucosamine by heparan sulfate 3-O-sulfotransferase-1 (3-OST-1) is akey modification step during the biosynthesis of anticoagulant heparan sulfate (HS). In this paper, wepresent evidence of a conformational change that occurs in 3-OST-1 upon binding to heparan sulfate.The intrinsic fluorescence of 3-OST-1 was increased in the presence of HS, suggesting a conformationalchange. This apparent conformational change was further investigated using differential chemicalmodification of 3-OST-1 to measure the solvent accessibility of the lysine residues. 3-OST-1 was treatedwith acetic anhydride in either the presence or absence of HS using both acetic anhydride andhexadeuterioacetic anhydride under nondenaturing and denaturing conditions, respectively. The relativereactivity of the lysine residues to acetylation and [2H] acetylation in the presence or absence of HS wasanalyzed by measuring the ratio of acetylated and deuterioacetylated peptides using matrix-assisted laserdesorption ionization mass spectrometry. The solvent accessibilities of the lysine residues were altereddifferentially depending on their location. In particular, we observed a group of lysine residues in theC-terminus of 3-OST-1 that become more solvent accessible when 3-OST-1 binds to HS. This observationindicates that a conformational change could be occurring during substrate binding. A truncated mutantof 3-OST-1 that lacked this C-terminal region was expressed and found to exhibit a 200-fold reductionin sulfotransferase activity. The results from this study will contribute to our understanding of theinteractions between 3-OSTs and HS.
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