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HEPES pH (HEPE + ph)
Selected AbstractsExpression, crystallization and preliminary X-ray crystallographic studies of Arthrobacter globiformis inulin fructotransferaseACTA CRYSTALLOGRAPHICA SECTION D, Issue 12 2003Mitsuru Momma A recombinant form of Arthrobacter globiformis inulin fructotransferase (DFAIII-producing) has been overexpressed in Escherichia coli and purified to homogeneity. Crystals were obtained at 293,K by the hanging-drop vapour-diffusion technique using 0.1,M Na HEPES pH 7.5 buffer containing 1.5,M lithium sulfate as a precipitant. Crystals of the recombinant wild-type enzyme diffracted to better than 1.5,Å at 100,K using a synchrotron-radiation source at the Photon Factory. The crystal belonged to space group R32, with unit-cell parameters a = b = 92.02, c = 229.82,Å in the hexagonal axes. Assuming the presence of one molecule in the asymmetric unit, the VM value for the crystal was 2.15,Å3,Da,1, indicating a solvent content of 42.8%. Selenomethionine-derivative crystals belonged to a different space group, C2, with unit-cell parameters a = 159.32, b = 91.92, c = 92.58,Å, , = 125.06. Matthews coefficient calculations suggested that the C2 selenomethionine-derivative crystal contained three molecules per asymmetric unit. [source] Crystallization and preliminary X-ray data of the recombinant peptide amidase from Stenotrophomonas maltophiliaACTA CRYSTALLOGRAPHICA SECTION D, Issue 2 2002Sebastian Neumann The peptide amidase from Stenotrophomonas maltophilia selectively hydrolyses the C-terminal amide bond in peptide amides. Crystals have been obtained by sitting-drop vapour diffusion from solution containing polyethylene glycol (PEG) 6000, HEPES pH 7.5, glycerine and sodium azide (NaN3). The crystals belong to the monoclinic space group P21, with unit-cell parameters a = 74.18, b = 62.60, c = 101.91,Å, , = 90°. X-ray data from these crystals diffracted at the European Synchrotron Radiation Facility (ESRF, France) ID14-1 beamline to 1.4,Å. [source] Structure of XynB, a highly thermostable ,-1,4-xylanase from Dictyoglomus thermophilum Rt46B.1, at 1.8,Å resolutionACTA CRYSTALLOGRAPHICA SECTION D, Issue 11 2000Andrew A. McCarthy Microorganisms employ a large array of enzymes to break down the cellulose and hemicelluloses of plant biomass. These enzymes, especially those with high thermal stability, have many uses in biotechnology. We have solved the crystal structure of a ,-1,4-xylanase, XynB, from the extremely thermophilic bacterium Dictyoglomus thermophilum, isolate Rt46B.1. The protein crystallized from 1.6,M ammonium sulfate, 0.2,M HEPES pH 7.2 and 10% glycerol, with unit-cell parameters a = b = 91.3, c = 44.9,Å and space group P43. The structure was solved at high resolution (1.8,Å) by X-ray crystallography, using the method of isomorphous replacement with a single mercury derivative, and refined to a final R factor of 18.3% (Rfree = 22.1%). XynB has the single-domain fold typical of family 11 xylanases, comprising a jelly roll of two highly twisted ,-sheets that create a deep substrate-binding cleft. The two catalytic residues, Glu90 and Glu180, occupy this cleft. Compared with other family 11 xylanases, XynB has a greater proportion of polar surface and has a slightly extended C-terminus that, combined with the extension of ,-strand A5, gives additional hydrogen bonding and hydrophobic packing. These factors may account for the enhanced thermal stability of the enzyme. [source] Preliminary crystallographic analysis of the N-terminal domain of FILIA, a protein essential for embryogenesisACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 9 2010Juke Wang FILIA is a component of the subcortical maternal complex that is essential for early stage embryogenesis. Its 6×His-tagged N-terminal domain was expressed in Escherichia coli and purified to homogeneity. Two types of crystals formed under different crystallization conditions during screening. Orthorhombic crystals appeared in a solution containing 1.4,M ammonium sulfate, 0.1,M Tris pH 8.2 and 12% glycerol, while tetragonal crystals were obtained using 15% PEG 4000 mixed with 0.1,M HEPES pH 7.5 and 15% 2-propanol. High-quality diffraction data were collected from the two crystal forms to resolutions of 1.8 and 2.2,Å, respectively, using synchrotron radiation. The Matthews coefficients indicated that the P212121 and P41212 crystals contained two molecules and one molecule per asymmetric unit, respectively. A selenomethionine-substituted sample failed to crystallize under the native conditions, but another orthorhombic crystal form was obtained under different conditions and anomalous diffraction data were collected. [source] Crystallization of the pneumococcal autolysin LytC: in-house phasing using novel lanthanide complexesACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 4 2010Inmaculada Pérez-Dorado LytC, one of the major autolysins from the human pathogen Streptococcus pneumoniae, has been crystallized as needles by the hanging-drop technique using 10%(w/v) PEG 3350 as precipitant and 10,mM HEPES pH 7.5. LytC crystals were quickly soaked in mother liquor containing 2,mM of the complex Gd-HPDO3A to produce derivatized crystals (LytCGd-HPDO3A). Both native LytC and isomorphous LytCGd-HPDO3A crystals were flash-cooled in a nitrogen flow at 120,K prior to X-ray data collection using an in-house Enraf,Nonius rotating-anode generator (, = 1.5418,Å) and a MAR345 imaging-plate detector. In both cases, good-quality diffraction patterns were obtained at high resolution. LytCGd-HPDO3A crystals allowed the collection of a SAD X-ray data set to 2.6,Å resolution indexed in terms of a P21 monoclinic unit cell with parameters a = 59.37, b = 67.16, c = 78.85,Å, , = 105.69°. The anomalous Patterson map allowed the identification of one heavy-atom binding site, which was sufficient for the calculation of an interpretable anomalous map at 2.6,Å resolution. [source] Cloning, overexpression, purification, crystallization and preliminary X-ray diffraction analysis of an atypical two-cysteine peroxiredoxin (SAOUHSC_01822) from Staphylococcus aureus NCTC 8325ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 11 2009Sudipta Bhattacharyya An atypical two-cysteine peroxidase, SAOUHSC_01822, from the virulent Staphylococcus aureus strain NCTC 8325 plays a major role in the reponse of the bacterium to oxidative stress. The protein was cloned, expressed, purified to homogeneity and crystallized. The protein was crystallized from 2,M ammonium sulfate, 0.1,M Na HEPES pH 7, 2%(v/v) PEG 400. A complete diffraction data set was collected to 2.3,Å resolution using a Rigaku MicroMax HF007 Cu,K, X-ray generator and a Rigaku R-AXIS IV++ detector. The crystals belonged to space group P21, with unit-cell parameters a = 43.50, b = 149.35, c = 73.73,Å, , = 104.4°, and contained four molecules in the asymmetric unit. [source] Crystallization and preliminary X-ray diffraction analysis of the lectin from Canavalia boliviana Piper seedsACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 3 2009Tales Rocha Moura Plant lectins are the most studied group of carbohydrate-binding proteins. Despite the high similarity between the members of the Diocleinae subtribe (Leguminosae) group, they present differing biological activities. Canavalia boliviana lectin (Cbol) was purified using a Sephadex G-50 column and crystallized in the presence of X-Man by hanging-drop vapour diffusion at 293,K. After optimization, crystals suitable for diffraction were obtained under the condition 0.1,M HEPES pH 7.5 and 3.0,M sodium formate. The crystal belonged to the monoclinic space group C2, with unit-cell parameters a = 126.70, b = 66.64, c = 64.99,Å, , = 90.0, , = 120.8, , = 90.0°. Assuming the presence of a dimer in the asymmetric unit, the solvent content was estimated to be about 46%. A complete data set was collected at 1.5,Å resolution. [source] | ||||||||||