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Type: Journal article
Title: Structural analysis and insights into the glycon specificity of the rice GH1 Os7BGlu26 β-D-mannosidase
Other Titles: Structural analysis and insights into the glycon specificity of the rice GH1 Os7BGlu26 beta-D-mannosidase
Author: Tankrathok, A.
Iglesias-Ferna, J.
Luang, S.
Robinson, R.
Kimura, A.
Rovira, C.
Hrmova, M.
Cairns, J.
Citation: Acta Crystallographica Section D-Biological Crystallography, 2013; 69(10):2124-2135
Publisher: Munksgaard Int Publ Ltd
Issue Date: 2013
ISSN: 0907-4449
Statement of
Anupong Tankrathok, Javier Iglesias-Fernández, Sukanya Luang, Robert C. Robinson, Atsuo Kimura, Carme Rovira, Maria Hrmova and James R. Ketudat Cairns
Abstract: Rice Os7BGlu26 is a GH1 family glycoside hydrolase with a threefold higher kcat/Km value for 4-nitrophenyl β-D-mannoside (4NPMan) compared with 4-nitrophenyl β-D-glucoside (4NPGlc). To investigate its selectivity for β-D-mannoside and β-D-glucoside substrates, the structures of apo Os7BGlu26 at a resolution of 2.20 Å and of Os7BGlu26 with mannose at a resolution of 2.45 Å were elucidated from isomorphous crystals in space group P212121. The (β/α)8-barrel structure is similar to other GH1 family structures, but with a narrower active-site cleft. The Os7BGlu26 structure with D-mannose corresponds to a product complex, with β-D-mannose in the (1)S5 skew-boat conformation. Docking of the (1)S3, (1)S5, (2)SO and (3)S1 pyranose-ring conformations of 4NPMan and 4NPGlc substrates into the active site of Os7BGlu26 indicated that the lowest energies were in the (1)S5 and (1)S3 skew-boat conformations. Comparison of these docked conformers with other rice GH1 structures revealed differences in the residues interacting with the catalytic acid/base between enzymes with and without β-D-mannosidase activity. The mutation of Tyr134 to Trp in Os7BGlu26 resulted in similar kcat/Km values for 4NPMan and 4NPGlc, while mutation of Tyr134 to Phe resulted in a 37-fold higher kcat/Km for 4NPMan than 4NPGlc. Mutation of Cys182 to Thr decreased both the activity and the selectivity for β-D-mannoside. It was concluded that interactions with the catalytic acid/base play a significant role in glycon selection.
Keywords: Glycoside Hydrolases; beta-Mannosidase; Crystallography, X-Ray; Mutagenesis, Site-Directed; Catalytic Domain; Protein Conformation; Substrate Specificity; Glycosylation; Hydrolysis; Oryza
Rights: © 2013 International Union of Crystallography
RMID: 0020132179
DOI: 10.1107/S0907444913020568
Appears in Collections:Agriculture, Food and Wine publications

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