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Title: Purification, crystallization and preliminary X-ray analysis of rice BGlu1 β-glucosidase with and without 2-deoxy-2-fluoro-β-D-glucoside
Authors: Jisnuson Svasti
Watchalee Chuenchor
Rodjana Opassiri
Salila Pengthaisong
Jirundon Yuvaniyama
James R. Ketudat Cairns
Suranaree University of Technology
Mahidol University
Keywords: Biochemistry, Genetics and Molecular Biology;Physics and Astronomy
Issue Date: 1-Aug-2006
Citation: Acta Crystallographica Section F: Structural Biology and Crystallization Communications. Vol.62, No.8 (2006), 798-801
Abstract: Rice (Oryza sativa) BGlu1 β-glucosidase was expressed in Escherichia coli with N-terminal thioredoxin and hexahistidine tags and purified by immobilized metal-affinity chromatography (IMAC). After removal of the N-terminal tags, cation-exchange and S-200 gel-filtration chromatography yielded a 50 kDa BGlu1 with >95% purity. The free enzyme and a complex with 2,4-dinitrophenyl-2-deoxy-2-fluoro-β-D-glucopyranoside inhibitor were crystallized by microbatch and hanging-drop vapour diffusion. Small tetragonal crystals of BGlu1 with and without inhibitor grew in 18%(w/v) PEG 8000 with 0.1 M sodium cacodylate pH 6.5 and 0.2 M zinc acetate. Crystals of BGlu1 with inhibitor were streak-seeded into 23%(w/v) PEG MME 5000, 0.2 M ammonium sulfate, 0.1 M MES pH 6.7 to yield larger crystals. Crystals with and without inhibitor diffracted to 2.15 and 2.75 Å resolution, respectively, and had isomorphous orthorhombic unit cells belonging to space group P2 12121. © 2006 International Union of Crystallography All rights reserved.
ISSN: 17443091
Appears in Collections:Scopus 2006-2010

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