purified recombinnat enzyme, hanging drop vapour diffusion method, micro-seeding, the protein crystallized in 1.6 M ammonium sulfate, 100 mM HEPES, pH 7.0, within 4 weeks, X-ray diffraction structure determination and analysis
purified wild-type enzyme, 0.002 ml of protein solution containing 4 mg/ml protein in 20 mM MES, pH 6.0, are mixed with 0.001 ml of reservoir solution containing 0.2 M ammonium sulphate, MES buffer, pH 6.5, and 30% w/v polyethyleneglycol-monomethylether 5500, 20°C, several days, X-ray diffraction structure determination and anaylsis at 2.7 A resolution, molecular replacement method, modeling
structures of Lin1840 wild-type and mutant D270A in complexes with sophorose and laminaribiose. Residue Arg572 forms many hydrogen bonds with sophorose and laminaribiose at subsite +1, a key factor for substrate selectivity. The opposite side of subsite +1 from Arg572 is connected to a large empty space, which may be subsite +2 for the binding of sophorotriose
free enzyme and its covalent intermediate with 2-deoxy-2-fluoroglucoside at 2.2 A and 1.55 A resolution, respectively. The complex with the 2-fluoroglucoside includes a glycerol molecule, which may make a nucleophilic attack on the anomeric carbon in a transglycosylation reaction. Sugars are positioned as acceptors for transglycosylation by their interactions with E176, the catalytic acid/base, and Y131. Residues I179, N190 and N245 appear to interact with the substrates
in complex with a covalent intermediate, 2-deoxy-2-fluoroglucoside and, n-octyl-beta-D-thioglucopyranoside, microbatch under oil method, using 0.1 M Bis-Tris, pH 6.5, and 20% PEG MME 5000
purified recombinant enzyme, free and in a complex with 2,4-dinitrophenyl-2-deoxy-2-fluoro-beta-D-glucopyranoside inhibitor, growth of small crystals in 18% w/v PEG 8000 with 0.1 M sodium cacodylate, pH 6.5, and 0.2 M zinc acetate, 1-2 days, microseeding into 23% w/v PEG MME 5000, 0.2 M ammonium sulfate, and 0.1 M MES, pH 6.7, resulting in larger crystals, 1-2 months, X-ray diffraction structure determination and analysis at 2.15 A and 2.75 A resolution, respectively, molecular replacement, screening and method optimization, overview
structures of the BGlu1 E176Q mutant, its complexes with cellotetraose, cellopentaose and laminaribiose, and its covalent intermediate with 2-deoxy-2-fluoroglucoside are determined at 1.65, 1.95, 1.80, 2.80, and 1.90 A resolution, respectively
thioredoxin/His6-tagged 4BGlu12 fusion protein complexed with 2,4-dinitrophenyl-2-deoxy-2-fluoro-beta-D-glucopyranoside, hanging drop vapor diffusion method, using 19% polyethylene glycol (3350 or 2000) in 0.1 M Tris-HCl pH 8.5, 0.16 M NaCl, at 15°C
isozyme BglB with bound inhibitors 2-deoxy-2-fluoro-alpha-D-glucopyranose or thiocellobiose, glucose, or as BglB-cellotetraose complex, X-ray difraction structure determination and analysis at 2.15-2.45 A resolution
isozyme BGL1A in substrate-free and gluconolactone complexed forms, sitting-drop vapor diffusion method at 25 °C, using a mixture of 0.001 ml of protein solution containing 10 mg/ml protein in 20 mM TrisHCl, pH 7.5, 150 mM NaCl, with or without 10 mM D-glucono-1,5-lactone, and 0.001 ml of a reservoir solution containing 12% isopropanol, 15% PEG 6000 and 0.1M sodium citrate, pH 5.8, the cryoprotectant solution contains 20% MPD, 15% PEG 6000 and 0.1M sodium citrate, pH 5.8, X-ray diffraction structure determination and analysis at 1.5-1.9 A resolution, modeling
mutant enzyme R170A/R220A/Y227F, hanging-drop vapour-diffusion method, the structure is determined at 2.35 A resolution, stable homotetrameric structure, a dimeric form of the enzyme is constructed and its crystal structure is solved at a resolution of 1.70 A
wild-type Dhr1, and Dhr1 mutant E189D in complex with dhurrin, hanging drop vapour diffusion method, 0.002 ml of 8 mg/ml protein in 20 mM Tris, pH 7.0, mixed with an equal volume of reservoir solution containing 0.1 M HEPES, pH 7.5, 2.0 M ammonium sulfate, and 50 mM NaCl, X-ray diffraction structure determination and analysis at 1.9-2.0 A resolution
at a resolution of 2.05A. It is composed of an (alpha/beta)8 domain similar to a triose phosphate isomerase barrel, a five-stranded alpha/beta sandwich domain (both of which are important for active-site organization), and a C-terminal fibronectin type III
purified recombinant wild-type and selenomethionine-substituted enzymes, hanging drop vapour diffusion method, 3-5 mg/ml protein in 20 mM MES buffer, pH 6.2, crystallization in microplates at 20°C within 4 days, precipitant is 90 mM bis-Tris propane buffer, pH 7.4, with 16-20% PEG 3350, 0.1-0.2 M NaI for the wild-type enzyme, and 20-24% PEG 3350, 0.1-0.25 M NaI for the selenomethionine-labeled enzyme, mixture of 0.001 ml of protein and reservoir solution, X-ray diffraction structure determination and analysis at 2.4 A resolution, MAD method, model building
purified recombinant enzyme complexed with a substrate aglycone, 20°C, hanging drop vapour diffusion method, 0.001 ml of 3.7 mg/ml protein in solution mixed with 0.001 ml reservoir solution containing 10 mM HEPES, pH 7.2, 1 M LiSO4, and 150 mM NaCl, X-ray diffraction structure determination and preliminary analysis at 1.7 A resolution, molecular replacement
in ligand-free form, and in complex form with D-glucose, with inhibitor isofagomine, and D-fucose, wild-type and mutants in residue N223, to 1.2-1.9 A resolution
molecular modeling of structure. Residues Asp333 and Glu530 may act as the catalytic nucleophile and acid/base, respectively. Residues Val87, Leu187, Thr368, and Trp474 may participate in substrate recognition and binding
to 2.75 A resolution. The predicted aglycone region is surrounded by hydrophobic residues Val220, Pro294, and Ala237. The residues His174 and Trp342 delimit the access to the -1 subsite and are critical to define the active site architecture
purified recombinant Glu1 mutant E191D in complex with cyclic inhibitor glucoetrazole, the crystallization solution is miced with 0.005 ml glycerol and 0.003 ml of inhibitor solution, X-ray diffraction structure determination and analysis at 1.95 A resolution
3.2.1.21 beta-glucosidase
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