Examining the substrate specificity of the enzyme 3-O-sulfotransferase that regulates the biosynthesis of The anticoagulant heparan sulfate

Heparan sulfate (HS) is a highly sulfated glycosaminoglycan (GAG) that affects a wide range of physiopathological functions. Its diverse functionality stems from the varying sulfation patterns along the polysaccharide chain. The current techniques for identifying the sequence of HS on the cell surface are limited. Thus, the goal of this project was to exploit the substrate specificity of the enzyme 3-O-sulfotransferase isoform 1 (3-OST 1) for the development of the HS binding probe that would recognize the anticoagulant HS. Thereby, molecular dynamics (MD) simulations and MM-GBSA analysis were performed to examine the structure and the free energy between 3-OST 1 and the 3-O-sulfated HS. In order to convert the 3-OST 1 into high-affinity binding probe, site-directed mutagenesis was performed on the catalytic glutamic acid (E90) residue and the effects of the mutations in binding to HS fragments were investigated by Bio-Layer Interferometry (BLI) analysis.