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Preliminary X-ray Crystallographic Study (preliminary + x-ray_crystallographic_study)
Selected AbstractsPreliminary X-ray crystallographic study of the receptor-binding domain of the D/C mosaic neurotoxin from Clostridium botulinumACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 5 2010Nipawan Nuemket Botulinum toxin (BoNT) from Clostridium botulinum OFD05, isolated from bovine botulism, is a D/C mosaic-type BoNT. BoNTs possess binding, translocation and catalytic domains. The BoNT/OFD05 binding domain exhibits significant sequence identity to BoNT/C, which requires a single ganglioside as a binding receptor on neuronal cells, while BoNT/A and BoNT/B require two receptors for specific binding. To determine the binding mechanism of BoNT/OFD05 and its ganglioside receptors on neuronal cells, recombinant BoNT/OFD05 receptor-binding domain has been expressed, purified and crystallized. Native and SeMet-derivative crystals showed X-ray diffraction to 2.8 and 3.1,Å resolution, respectively. The crystals belonged to space group P212121. [source] Crystallization and preliminary X-ray crystallographic study of leucyl-tRNA synthetase from the archaeon Pyrococcus horikoshiiACTA CRYSTALLOGRAPHICA SECTION D, Issue 10 2004Ryuya Fukunaga The leucyl-tRNA synthetase (LeuRS) from the archaeon Pyrococcus horikoshii was overexpressed in a C-terminally truncated form in Escherichia coli, purified and crystallized by the hanging-drop vapour-diffusion method using ammonium sulfate as a precipitant. The crystals belong to the rhombohedral space group R3, with unit-cell parameters a = b = 186.20, c = 91.43,Å, , = , = 90, , = 120°. The asymmetric unit contains one molecule of LeuRS, with a corresponding crystal volume per protein weight of 3.2,Å3,Da,1 and a solvent content of 60.7%. A data set diffracting to 2.2,Å resolution was collected from a single crystal at 100,K. Selenomethionine-substituted protein crystals were prepared in order to solve the structure by the SAD phasing method. [source] Crystallization and preliminary X-ray crystallographic study on a xyloglucan-specific exo-,-glycosidase, oligoxyloglucan reducing-end specific cellobiohydrolaseACTA CRYSTALLOGRAPHICA SECTION D, Issue 10 2003Katsuro Yaoi A novel xyloglucan-specific exo-,-glycosidase, oligoxyloglucan reducing-end specific cellobiohydrolase (OXG-RCBH), recognizes the reducing end of oligoxyloglucan and releases two glucosyl residue segments from the main chain. OXG-RCBH was crystallized by the hanging-drop vapour-diffusion method with polyethylene glycol 3000 and polyethylene glycol 400,as precipitants. The crystals belong to the orthorhombic space group P212121, with unit-cell parameters a = 61.0, b = 146.9, c = 211.9,Å. The crystals diffract to a resolution of 2.2,Å and are suitable for X-ray structure analysis. [source] Crystallization and preliminary X-ray crystallographic study of GenX, a lysyl-tRNA synthetase paralogue from Escherichia coli, in complex with translation elongation factor PACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 9 2010Tomomi Sumida GenX, a lysyl-tRNA synthetase paralogue from Escherichia coli, was overexpressed in E. coli, purified by three chromatographic steps and cocrystallized with a lysyl adenylate analogue (LysAMS) by the hanging-drop vapour-diffusion method using PEG 4000 as a precipitant. The GenX,LysAMS crystals belonged to the triclinic space group P1, with unit-cell parameters a = 54.80, b = 69.15, c = 94.08,Å, , = 95.47, , = 106.51, , = 90.46°, and diffracted to 1.9,Å resolution. Furthermore, GenX was cocrystallized with translation elongation factor P (EF-P), which is believed to be a putative substrate of GenX, and LysAMS using PEG 4000 and ammonium sulfate as precipitants. The GenX,EF-P,LysAMS crystals belonged to the monoclinic space group P21, with unit-cell parameters a = 105.93, b = 102.96, c = 119.94,Å, , = 99.4°, and diffracted to 2.5,Å resolution. Structure determination of the E. coli GenX,LysAMS and GenX,EF-P,LysAMS complexes by molecular replacement was successful and structure refinements are now in progress. [source] Expression, crystallization and preliminary X-ray crystallographic study of ethanolamine ammonia-lyase from Escherichia coliACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 6 2010Naoki Shibata Ethanolamine ammonia-lyase (EAL) catalyzes the adenosylcobalamin-dependent conversion of ethanolamine to acetaldehyde and ammonia. The wild-type enzyme shows a very low solubility. N-terminal truncation of the Escherichia coli EAL ,-subunit dramatically increases the solubility of the enzyme without altering its catalytic properties. Two deletion mutants of the enzyme [EAL(,,4,30) and EAL(,,4,43)] have been overexpressed, purified and crystallized using the sitting-drop vapour-diffusion method. Crystals of EAL(,,4,30) and EAL(,,4,43) diffracted to approximately 8.0 and 2.1,Å resolution, respectively. [source] Crystallization and preliminary X-ray crystallographic study of 1-deoxy- d -xylulose 5-phosphate reductoisomerase from Plasmodium falciparumACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 3 2010Tomonobu Umeda The nonmevalonate pathway of isoprenoid biosynthesis present in Plasmodium falciparum is known to be an effective target for antimalarial drugs. The second enzyme of the nonmevalonate pathway, 1-deoxy- d -xylulose 5-phosphate reductoisomerase (DXR), catalyzes the transformation of 1-deoxy- d -xylulose 5-phosphate (DXP) to 2- C -methyl- d -erythritol 4-phosphate (MEP). For crystallographic studies, DXR from the human malaria parasite P. falciparum (PfDXR) was overproduced in Escherichia coli, purified and crystallized using the hanging-drop vapour-diffusion method in the presence of NADPH. X-ray diffraction data to 1.85,Å resolution were collected from a monoclinic crystal form belonging to space group C2 with unit-cell parameters a = 168.89, b = 59.65, c = 86.58,Å, , = 117.8°. Structural analysis by molecular replacement is in progress. [source] Crystallization and preliminary X-ray crystallographic study of phosphoglucose isomerase from Plasmodium falciparumACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 3 2010Ken-ichi Aoki Phosphoglucose isomerase (PGI) is a key enzyme in glycolysis and glycogenesis that catalyses the interconversion of glucose 6-phosphate (G6P) and fructose 6-phosphate (F6P). For crystallographic studies, PGI from the human malaria parasite Plasmodium falciparum (PfPGI) was overproduced in Escherichia coli, purified and crystallized using the hanging-drop vapour-diffusion method. X-ray diffraction data to 1.5,Å resolution were collected from an orthorhombic crystal form belonging to space group P212121 with unit-cell parameters a = 103.3, b = 104.1, c = 114.6,Å. Structural analysis by molecular replacement is in progress. [source] Cloning, expression, purification, crystallization and preliminary X-ray crystallographic study of the putative SAICAR synthetase (PH0239) from Pyrococcus horikoshii OT3ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 2 2010Kavyashree Manjunath The study of proteins involved in de novo biosynthesis of purine nucleotides is central in the development of antibiotics and anticancer drugs. In view of this, a protein from the hyperthermophile Pyrococcus horikoshii OT3 was isolated, purified and crystallized using the microbatch method. Its primary structure was found to be similar to that of SAICAR synthetase, which catalyses the seventh step of de novo purine biosynthesis. A diffraction-quality crystal was obtained using Hampton Research Crystal Screen II condition No. 34, consisting of 0.05,M cadmium sulfate hydrate, 0.1,M HEPES buffer pH 7.5 and 1.0,M sodium acetate trihydrate, with 40%(v/v) 1,4-butanediol as an additive. The crystal belonged to space group P31, with unit-cell parameters a = b = 95.62, c = 149.13,Å. Assuming the presence of a hexamer in the asymmetric unit resulted in a Matthews coefficient (VM) of 2.3,Å3,Da,1, corresponding to a solvent content of about 46%. A detailed study of this protein will yield insights into structural stability at high temperatures and should be highly relevant to the development of antibiotics and anticancer drugs targeting the biosynthesis of purine nucleotides. [source] Expression and purification of F-plasmid RepE and preliminary X-ray crystallographic study of its complex with operator DNAACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 4 2007Chieko Wada The replication initiator factor RepE of the F plasmid in Escherichia coli is an essential protein that stringently regulates the F-plasmid copy number. The RepE protein has a dual function: its monomer functions as a replication initiator, while its dimer acts as a transcriptional repressor of the repE gene. The wild-type dimeric RepE protein was expressed as an N-terminal histidine-tagged protein, purified under native conditions with a high salt concentration and crystallized in complex with the repE operator DNA using the sitting-drop vapour-diffusion technique. The crystals diffracted to a resolution of 3.14,Å after the application of dehydration and crystal annealing and belong to space group P21, with unit-cell parameters a = 60.73, b = 99.32, c = 95.00,Å, , = 108.55°. [source] Purification, crystallization and preliminary X-ray crystallographic study of the l -fuculose-1-phosphate aldolase (FucA) from Thermus thermophilus HB8ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 12 2005Jeyaraman Jeyakanthan Fuculose phosphate aldolase catalyzes the reversible cleavage of l -fuculose-1-phosphate to dihydroxyacetone phosphate and l -lactaldehyde. The protein from Thermus thermophilus HB8 is a biological tetramer with a subunit molecular weight of 21,591,Da. Purified FucA has been crystallized using sitting-drop vapour-diffusion and microbatch techniques at 293,K. The crystals belong to space group P4, with unit-cell parameters a = b = 100.94, c = 45.87,Å. The presence of a dimer of the enzyme in the asymmetric unit was estimated to give a Matthews coefficient (VM) of 2.7,Å3,Da,1 and a solvent content of 54.2%(v/v). Three-wavelength diffraction MAD data were collected to 2.3,Å from zinc-containing crystals. Native diffraction data to 1.9,Å resolution have been collected using synchrotron radiation at SPring-8. [source] Crystallization and preliminary X-ray crystallographic study of disproportionating enzyme from potatoACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 1 2005Kayo Imamura Disproportionating enzyme (D-enzyme; EC 2.4.1.25) is a 59,kDa protein that belongs to the ,-amylase family. D-enzyme catalyses intramolecular and intermolecular transglycosylation reactions of ,-1,4 glucan. A crystal of the D-enzyme from potato was obtained by the hanging-drop vapour-diffusion method. Preliminary X-ray data showed that the crystal diffracts to 2.0,Å resolution and belongs to space group C2221, with unit-cell parameters a = 69.7, b = 120.3, c = 174.2,Å. [source] | ||||||||||