Structural determinants of substrate recognition and catalysis by heparan sulfate sulfotransferases

Tarsis F Gesteira, Tainah Marforio, Jonathan Wolf Mueller, Matteo Calvaresi, Vivien Jane Coulson-Thomas

Research output: Contribution to journalArticlepeer-review

Abstract

Heparan sulfate (HS) and heparin contain imprinted “sulfation codes”, which dictate their diverse physiological and pathological functions. A group of orchestrated biosynthetic enzymes cooperate in polymerizing and modifying HS chains. The biotechnological development of enzymes that can recreate this sulfation pattern on synthetic heparin is challenging, primarily due to the paucity of quantitative data for sulfotransferase enzymes. Herein, we identified critical structural characteristics that determine substrate specificity and shed light on the catalytic mechanism of sugar sulfation of two HS sulfotransferases, 2-O-sulfotransferase (HS2ST) and 6-O-sulfotransferase (HS6ST). Two sets of molecular clamps in HS2ST recognize appropriate substrates; these clamps flank the acceptor binding site on opposite sides. The hexuronic epimers, and not their puckers, have a critical influence on HS2ST selectivity. In contrast, HS6ST recognizes a broader range of substrates. This promiscuity is granted by a conserved tryptophan residue, W210, that positions the acceptor within the active site for catalysis by means of strong electrostatic interactions. Lysines K131 and K132 act in concert with a second tryptophan, W153, shedding water molecules from within the active site, thus providing HS6ST with a binding preference toward 2-O-sulfated substrates. QM/MM calculations provided valuable mechanistic insights into the catalytic process, identifying that the sulfation of both HS2ST and HS6ST follows a SN2-like mechanism. When they are taken together, our findings reveal the molecular basis of how these enzymes recognize different substrates and catalyze sugar sulfation, enabling the generation of enzymes that could create specific heparin epitopes.
Original languageEnglish
Pages (from-to)10974–10987
Number of pages14
JournalACS Catalysis
Volume11
Issue number17
Early online date18 Aug 2021
DOIs
Publication statusPublished - 3 Sep 2021

Keywords

  • heparin
  • glycosaminoglycan biosynthesis
  • sulfotransferase
  • sulfation
  • HS2ST
  • HS6ST
  • sugar ring puckering
  • active-site solvation

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