title: Chemoenzymatic synthesis of 6-phospho-cyclophellitol as a novel probe of 6-phospho-β-glucosidases.
abstract: Covalent, mechanism-based inhibitors of glycosidases are valuable probe molecules for visualizing enzyme activities in complex systems. We, here, describe the chemoenzymatic synthesis of 6-phospho-cyclophellitol and evaluate its behaviour as a mechanism-based inactivator of the Streptococcus pyogenes 6-phospho-β-glucosidase from CAZy family GH1. We further present the three-dimensional structure of the inactivated enzyme, which reveals the constellation of active site residues responsible for the enzyme's specificity and confirms the covalent nature of the inactivation.
authors.affiliation: Department of Chemistry, University of British Columbia, BC, Canada.
authors.forename: David H
authors.initials: DH
authors.lastname: Kwan
authors.affiliation: York Structural Biology Laboratory, Department of Chemistry, University of York, UK.
authors.forename: Yi
authors.initials: Y
authors.lastname: Jin
authors.affiliation: Department of Bioorganic Synthesis, Leiden Institute of Chemistry, Leiden University, the Netherlands.
authors.forename: Jianbing
authors.initials: J
authors.lastname: Jiang
authors.affiliation: Department of Chemistry, University of British Columbia, BC, Canada.
authors.forename: Hong-Ming
authors.initials: HM
authors.lastname: Chen
authors.affiliation: Department of Chemistry, University of British Columbia, BC, Canada.
authors.forename: Miriam P
authors.initials: MP
authors.lastname: Kötzler
authors.affiliation: Department of Bioorganic Synthesis, Leiden Institute of Chemistry, Leiden University, the Netherlands.
authors.forename: Herman S
authors.initials: HS
authors.lastname: Overkleeft
authors.affiliation: York Structural Biology Laboratory, Department of Chemistry, University of York, UK.
authors.forename: Gideon J
authors.initials: GJ
authors.lastname: Davies
authors.affiliation: Department of Chemistry, University of British Columbia, BC, Canada.
authors.forename: Stephen G
authors.initials: SG
authors.lastname: Withers
mesh_terms: D020134:Catalytic Domain; D003067:Coenzymes / Q000737:chemistry; D018360:Crystallography, X-Ray; D003511:Cyclohexanols / Q000138:chemical synthesis* / Q000737:chemistry; D004791:Enzyme Inhibitors / Q000138:chemical synthesis* / Q000737:chemistry; D005959:Glucosidases / Q000037:antagonists & inhibitors* / Q000737:chemistry; D015335:Molecular Probes / Q000138:chemical synthesis* / Q000737:chemistry; D011487:Protein Conformation; D013297:Streptococcus pyogenes / Q000201:enzymology*
chemical_list: D003067:Coenzymes; D003511:Cyclohexanols; D004791:Enzyme Inhibitors; D015335:Molecular Probes; C063285:cyclophellitol; D005959:Glucosidases; C066586:6-phospho-beta-glucosidase
keywords: CAZy GH1; Glycosidase inhibitors; activity-based probes; epoxides; sugar-6-phosphate