The structural basis of oligosaccharide binding by rice BGlu1 beta-glucosidase

Watchalee Chuenchor, Salila Pengthaisong, Robert C. Robinson, Jirundon Yuvaniyama, Jisnuson Svasti, James R.Ketudat Cairns

Research output: Contribution to journalArticle

34 Citations (Scopus)

Abstract

Rice BGlu1 β-glucosidase is an oligosaccharide exoglucosidase that binds to six β-(1→4)-linked glucosyl residues in its active site cleft. Here, we demonstrate that a BGlu1 E176Q active site mutant can be effectively rescued by small nucleophiles, such as acetate, azide and ascorbate, for hydrolysis of aryl glycosides in a pH-independent manner above pH. 5, consistent with the role of E176 as the catalytic acid-base. Cellotriose, cellotetraose, cellopentaose, cellohexaose and laminaribiose are not hydrolyzed by the mutant and instead exhibit competitive inhibition. The structures of the BGlu1 E176Q, its complexes with cellotetraose, cellopentaose and laminaribiose, and its covalent intermediate with 2-deoxy-2-fluoroglucoside were determined at 1.65, 1.95, 1.80, 2.80, and 1.90. Å resolution, respectively. The Q176. Nε was found to hydrogen bond to the glycosidic oxygen of the scissile bond, thereby explaining its high activity. The enzyme interacts with cellooligosaccharides through direct hydrogen bonds to the nonreducing terminal glucosyl residue. However, interaction with the other glucosyl residues is predominantly mediated through water molecules, with the exception of a direct hydrogen bond from N245 to glucosyl residue 3, consistent with the apparent high binding energy at this residue. Hydrophobic interactions with the aromatic sidechain of W358 appear to orient glucosyl residues 2 and 3, while Y341 orients glucosyl residues 4 and 5. In contrast, laminaribiose has its second glucosyl residue positioned to allow direct hydrogen bonding between its O2 and Q176 Oε and O1 and N245. These are the first GH1 glycoside hydrolase family structures to show oligosaccharide binding in the hydrolytic configuration.

Original languageEnglish
Pages (from-to)169-179
Number of pages11
JournalJournal of Structural Biology
Volume173
Issue number1
DOIs
Publication statusPublished - Jan 1 2011
Externally publishedYes

Fingerprint

beta-Glucosidase
Oligosaccharides
Hydrogen
Catalytic Domain
Glucosidases
Azides
Glycoside Hydrolases
Hydrogen Bonding
Glycosides
Hydrophobic and Hydrophilic Interactions
Acetates
Hydrolysis
Oxygen
Acids
Water
Enzymes
laminaribiose
Oryza
cellotetraose
maltopentaose

Keywords

  • Acid-base mutant
  • Cello-oligosaccharides
  • Enzyme-substrate complex
  • Glycoside hydrolase
  • X-ray crystallography

ASJC Scopus subject areas

  • Structural Biology

Cite this

The structural basis of oligosaccharide binding by rice BGlu1 beta-glucosidase. / Chuenchor, Watchalee; Pengthaisong, Salila; Robinson, Robert C.; Yuvaniyama, Jirundon; Svasti, Jisnuson; Cairns, James R.Ketudat.

In: Journal of Structural Biology, Vol. 173, No. 1, 01.01.2011, p. 169-179.

Research output: Contribution to journalArticle

Chuenchor, W, Pengthaisong, S, Robinson, RC, Yuvaniyama, J, Svasti, J & Cairns, JRK 2011, 'The structural basis of oligosaccharide binding by rice BGlu1 beta-glucosidase', Journal of Structural Biology, vol. 173, no. 1, pp. 169-179. https://doi.org/10.1016/j.jsb.2010.09.021
Chuenchor, Watchalee ; Pengthaisong, Salila ; Robinson, Robert C. ; Yuvaniyama, Jirundon ; Svasti, Jisnuson ; Cairns, James R.Ketudat. / The structural basis of oligosaccharide binding by rice BGlu1 beta-glucosidase. In: Journal of Structural Biology. 2011 ; Vol. 173, No. 1. pp. 169-179.
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