A Single Glycosidase Harnesses Different Pyranoside Ring Transition State Conformations for Hydrolysis of Mannosides and Glucosides

Anupong Tankrathok, Javier Iglesias-Fernández, Rohan J. Williams, Salila Pengthaisong, Supaporn Baiya, Zalihe Hakki, Robert C. Robinson, Maria Hrmova, Carme Rovira, Spencer J. Williams, James R. Ketudat Cairns

Research output: Contribution to journalArticle

11 Citations (Scopus)


Hydrolysis of β-d-mannosides by β-mannosidases typically proceeds via a B2,5 transition state conformation for the pyranoside ring, while that of β-d-glucosides by β-glucosidases proceeds through a distinct 4H3 transition state conformation. However, rice Os7BGlu26 β-glycosidase hydrolyzes 4-nitrophenyl β-d-glucoside and β-d-mannoside with similar efficiencies. The origin of this dual substrate specificity was investigated by kinetic, structural, and computational approaches. The glycosidase inhibitors glucoimidazole and mannoimidazole inhibited Os7BGlu26 with Ki values of 2.7 nM and 10.4 μM, respectively. In X-ray crystal structures of complexes with Os7BGlu26, glucoimidazole bound to the active site in a 4E conformation, while mannoimidazole bound in a B2,5 conformation, suggesting different transition state conformations. Moreover, calculation of quantum mechanics/molecular mechanics (QM/MM) free energy landscapes showed that 4-nitrophenyl β-d-glucoside adopts a 1S3/4E conformation in the Michaelis complex, while 4-nitrophenyl β-d-mannoside adopts a 1S5/B2,5 conformation. The QM/MM simulations of Os7BGlu26 catalysis of hydrolysis also supported the itineraries of 1S34E/4H34C1 for β-d-glucosides and 1S5 → B2,5OS2 for β-d-mannosides, thereby revealing that a single glycoside hydrolase can hydrolyze glycosides of different configurations via distinct transition state pyranoside conformations.

Original languageEnglish
Pages (from-to)6041-6051
Number of pages11
JournalACS Catalysis
Issue number10
Publication statusPublished - Oct 2 2015
Externally publishedYes



  • QM/MM metadynamics
  • structural analysis
  • transition state mimic
  • X-ray crystallography
  • β-glucosidase
  • β-mannosidase

ASJC Scopus subject areas

  • Catalysis
  • Chemistry(all)

Cite this

Tankrathok, A., Iglesias-Fernández, J., Williams, R. J., Pengthaisong, S., Baiya, S., Hakki, Z., Robinson, R. C., Hrmova, M., Rovira, C., Williams, S. J., & Ketudat Cairns, J. R. (2015). A Single Glycosidase Harnesses Different Pyranoside Ring Transition State Conformations for Hydrolysis of Mannosides and Glucosides. ACS Catalysis, 5(10), 6041-6051. https://doi.org/10.1021/acscatal.5b01547