An Epoxide Intermediate in Glycosidase Catalysis.

Autor: Sobala LF; York Structural Biology Laboratory, Department of Chemistry, University of York, York YO10 5DD, United Kingdom., Speciale G; School of Chemistry and Bio21 Molecular Science and Biotechnology Institute, University of Melbourne, Parkville, Victoria 3010, Australia., Zhu S; Sorbonne Université, CNRS, Institut Parisien de Chimie Moléculaire, UMR 8232, 4 place Jussieu, 75005 Paris, France., Raich L; Departament de Química Inorgànica i Orgànica (Secció de Química Orgànica) & Institut de Química Teòrica i Computacional (IQTCUB), Universitat de Barcelona, Martí i Franquès 1, 08028 Barcelona, Spain., Sannikova N; Department of Chemistry, Simon Fraser University, 8888 University Drive, Burnaby, British Columbia V5A 1S6, Canada., Thompson AJ; York Structural Biology Laboratory, Department of Chemistry, University of York, York YO10 5DD, United Kingdom., Hakki Z; School of Chemistry and Bio21 Molecular Science and Biotechnology Institute, University of Melbourne, Parkville, Victoria 3010, Australia., Lu D; Sorbonne Université, CNRS, Institut Parisien de Chimie Moléculaire, UMR 8232, 4 place Jussieu, 75005 Paris, France., Shamsi Kazem Abadi S; Department of Biochemistry and Molecular Biology, Simon Fraser University, 8888 University Drive, Burnaby, British Columbia V5A 1S6, Canada., Lewis AR; Department of Chemistry, Simon Fraser University, 8888 University Drive, Burnaby, British Columbia V5A 1S6, Canada., Rojas-Cervellera V; Departament de Química Inorgànica i Orgànica (Secció de Química Orgànica) & Institut de Química Teòrica i Computacional (IQTCUB), Universitat de Barcelona, Martí i Franquès 1, 08028 Barcelona, Spain., Bernardo-Seisdedos G; Molecular Recognition and Host-Pathogen Interactions, CIC bioGUNE, Basque Research Technology Alliance (BRTA), Bizkaia Technology Park, Building 800, 48160 Derio, Spain., Zhang Y; Sorbonne Université, CNRS, Institut Parisien de Chimie Moléculaire, UMR 8232, 4 place Jussieu, 75005 Paris, France., Millet O; Molecular Recognition and Host-Pathogen Interactions, CIC bioGUNE, Basque Research Technology Alliance (BRTA), Bizkaia Technology Park, Building 800, 48160 Derio, Spain., Jiménez-Barbero J; Ikerbasque, Basque Foundation for Science, Marıá Dıáz de Haro 3, 48013 Bilbao, Spain.; Molecular Recognition and Host-Pathogen Interactions, CIC bioGUNE, Basque Research Technology Alliance (BRTA), Bizkaia Technology Park, Building 800, 48160 Derio, Spain., Bennet AJ; Department of Chemistry, Simon Fraser University, 8888 University Drive, Burnaby, British Columbia V5A 1S6, Canada.; Department of Biochemistry and Molecular Biology, Simon Fraser University, 8888 University Drive, Burnaby, British Columbia V5A 1S6, Canada., Sollogoub M; Sorbonne Université, CNRS, Institut Parisien de Chimie Moléculaire, UMR 8232, 4 place Jussieu, 75005 Paris, France., Rovira C; Departament de Química Inorgànica i Orgànica (Secció de Química Orgànica) & Institut de Química Teòrica i Computacional (IQTCUB), Universitat de Barcelona, Martí i Franquès 1, 08028 Barcelona, Spain.; Institució Catalana de Recerca i Estudis Avançats (ICREA), Pg. Lluís Companys 23, 08010 Barcelona, Spain., Davies GJ; York Structural Biology Laboratory, Department of Chemistry, University of York, York YO10 5DD, United Kingdom., Williams SJ; School of Chemistry and Bio21 Molecular Science and Biotechnology Institute, University of Melbourne, Parkville, Victoria 3010, Australia.
Jazyk: angličtina
Zdroj: ACS central science [ACS Cent Sci] 2020 May 27; Vol. 6 (5), pp. 760-770. Date of Electronic Publication: 2020 Apr 16.
DOI: 10.1021/acscentsci.0c00111
Abstrakt: Retaining glycoside hydrolases cleave their substrates through stereochemical retention at the anomeric position. Typically, this involves two-step mechanisms using either an enzymatic nucleophile via a covalent glycosyl enzyme intermediate or neighboring-group participation by a substrate-borne 2-acetamido neighboring group via an oxazoline intermediate; no enzymatic mechanism with participation of the sugar 2-hydroxyl has been reported. Here, we detail structural, computational, and kinetic evidence for neighboring-group participation by a mannose 2-hydroxyl in glycoside hydrolase family 99 endo -α-1,2-mannanases. We present a series of crystallographic snapshots of key species along the reaction coordinate: a Michaelis complex with a tetrasaccharide substrate; complexes with intermediate mimics, a sugar-shaped cyclitol β-1,2-aziridine and β-1,2-epoxide; and a product complex. The 1,2-epoxide intermediate mimic displayed hydrolytic and transfer reactivity analogous to that expected for the 1,2-anhydro sugar intermediate supporting its catalytic equivalence. Quantum mechanics/molecular mechanics modeling of the reaction coordinate predicted a reaction pathway through a 1,2-anhydro sugar via a transition state in an unusual flattened, envelope ( E 3 ) conformation. Kinetic isotope effects ( k cat / K M ) for anomeric- 2 H and anomeric- 13 C support an oxocarbenium ion-like transition state, and that for C2- 18 O (1.052 ± 0.006) directly implicates nucleophilic participation by the C2-hydroxyl. Collectively, these data substantiate this unprecedented and long-imagined enzymatic mechanism.
Competing Interests: The authors declare no competing financial interest.
(Copyright © 2020 American Chemical Society.)
Databáze: MEDLINE
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