Structural insights into choline-O-sulfatase reveal the molecular determinants for ligand binding

Acta Crystallogr D Struct Biol. 2022 May 1;78(Pt 5):669-682. doi: 10.1107/S2059798322003709. Epub 2022 Apr 26.


Choline-O-sulfatase (COSe; EC is a member of the alkaline phosphatase (AP) superfamily, and its natural function is to hydrolyze choline-O-sulfate into choline and sulfate. Despite its natural function, the major interest in this enzyme resides in the landmark catalytic/substrate promiscuity of sulfatases, which has led to attention in the biotechnological field due to their potential in protein engineering. In this work, an in-depth structural analysis of wild-type Sinorhizobium (Ensifer) meliloti COSe (SmeCOSe) and its C54S active-site mutant is reported. The binding mode of this AP superfamily member to both products of the reaction (sulfate and choline) and to a substrate-like compound are shown for the first time. The structures further confirm the importance of the C-terminal extension of the enzyme in becoming part of the active site and participating in enzyme activity through dynamic intra-subunit and inter-subunit hydrogen bonds (Asn146A-Asp500B-Asn498B). These residues act as the `gatekeeper' responsible for the open/closed conformations of the enzyme, in addition to assisting in ligand binding through the rearrangement of Leu499 (with a movement of approximately 5 Å). Trp129 and His145 clamp the quaternary ammonium moiety of choline and also connect the catalytic cleft to the C-terminus of an adjacent protomer. The structural information reported here contrasts with the proposed role of conformational dynamics in promoting the enzymatic catalytic proficiency of an enzyme.

Keywords: alkaline phosphatases; choline; conformational gating; promiscuity; sulfatases.

MeSH terms

  • Alkaline Phosphatase / chemistry
  • Alkaline Phosphatase / metabolism
  • Binding Sites
  • Choline
  • Ligands
  • Sinorhizobium meliloti*
  • Substrate Specificity
  • Sulfatases* / chemistry
  • Sulfatases* / metabolism
  • Sulfates


  • Ligands
  • Sulfates
  • Alkaline Phosphatase
  • Sulfatases
  • Choline