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Orthorhombic Crystals (orthorhombic + crystal)
Terms modified by Orthorhombic Crystals Selected AbstractsCloning, expression, purification and crystallization of a transcriptional regulatory protein (Rv3291c) from Mycobacterium tuberculosis H37RvACTA CRYSTALLOGRAPHICA SECTION D, Issue 10 2004Tripti Shrivastava Rv3291c, the translational product of the Mycobacterium tuberculosisRv3291c gene, is an 18,kDa protein. It is a putative transcriptional regulatory protein belonging to the leucine-responsive regulatory protein/asparagine synthase C (Lrp/AsnC) family, which are proteins that have been identified in archaea and bacteria. Rv3291c probably plays a significant role during the persistent/latent phase of M. tuberculosis, as supported by its up-regulation several-fold during this stage. Orthorhombic crystals of recombinant Rv3291c have been grown from trisodium citrate dihydrate-buffered solutions containing monoammonium dihydrogen phosphate. Diffraction data extending to 2.7,Å have been collected from a single crystal with unit-cell parameters a = 99.6, b = 100.7, c = 100.6,Å. Assuming an octamer in the asymmetric unit results in a Matthews coefficient (VM) of 1.75,Å3,Da,1, corresponding to a solvent content of about 30%. [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] Structure of the T109S mutant of Escherichia coli dihydroorotase complexed with the inhibitor 5-fluoroorotate: catalytic activity is reflected by the crystal formACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 3 2007Mihwa Lee Crystals of a single-point mutant (T109S) of Escherichia coli dihydroorotase (DHOase) with diminished activity grown in the presence of l -dihydroorotate (l -DHO) are tetragonal, with a monomer in the asymmetric unit. These crystals are extremely unstable and disintegrate shortly after formation, which is followed by the growth of orthorhombic crystals from the remnants of the tetragonal crystals or at new nucleation sites. Orthorhombic crystals, for which a structure has previously been reported [Thoden et al. (2001), Biochemistry, 40, 6989,6997; Lee et al. (2005), J. Mol. Biol.348, 523,533], contain a dimer of DHOase in the asymmetric unit; the active site of one monomer contains the substrate N -carbamyl- l -aspartate (l -CA-asp) and the active site of the other monomer contains the product of the reaction, l -DHO. In the subunit with l -DHO in the active site, a surface loop (residues 105,115) is `open'. In the other subunit, with l -CA-asp in the active site, the loop folds inwards, forming specific hydrogen bonds from the loop to the l -CA-asp. The tetragonal crystal form can be stabilized by crystallization in the presence of the inhibitor 5-fluoroorotate (FOA), a product (l -DHO) mimic. Crystals of the complex of T109S DHOase with FOA are tetragonal, space group P41212, with unit-cell parameters a = b = 72.6, c = 176.1,Å. The structure has been refined to R and Rfree values of 0.218 and 0.257, despite severe anisotropy of the diffraction. In this structure, the flexible loops are both in the `open' conformation, which is consistent with FOA, like l -DHO, binding at both sites. The behaviour of the T109S mutant crystals of DHOase in the presence of l -DHO is explained by initial binding of l -DHO to both subunits, followed by slow conversion to l -CA-asp, with consequent movement of the flexible loop and dissolution of the crystals. Orthorhombic crystals are then able to grow in the presence of l -DHO and l -CA-asp. [source] Preliminary X-ray analysis of twinned crystals of sarcosine dimethylglycine methyltransferase from Halorhodospira halochorisACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 8 2009Juha Pekka Kallio Sarcosine dimethylglycine methyltransferase (EC 2.1.1.157) is an enzyme from the extremely halophilic anaerobic bacterium Halorhodospira halochoris. This enzyme catalyzes the twofold methylation of sarcosine to betaine, with S -adenosylmethionine (AdoMet) as the methyl-group donor. This study presents the crystallization and preliminary X-ray analysis of recombinant sarcosine dimethylglycine methyltransferase produced in Escherichia coli. Mass spectroscopy was used to determine the purity and homogeneity of the enzyme material. Two different crystal forms, which initially appeared to be hexagonal and tetragonal, were obtained. However, on analyzing the diffraction data it was discovered that both crystal forms were pseudo-merohedrally twinned. The true crystal systems were monoclinic and orthorhombic. The monoclinic crystal diffracted to a maximum of 2.15,Å resolution and the orthorhombic crystal diffracted to 1.8,Å resolution. [source] Crystallization and preliminary X-ray diffraction studies of AsaP1_E294A and AsaP1_E294Q, two inactive mutants of the toxic zinc metallopeptidase AsaP1 from Aeromonas salmonicida subsp. achromogenesACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 7 2009Xenia Bogdanovi Two mutants of the toxic extracellular zinc endopeptidase AsaP1 (AsaP1_E294Q and AsaP1_E294A) of Aeromonas salmonicida subsp. achromogenes were expressed in Escherichia coli and crystallized by the vapour-diffusion method. Crystals were obtained using several precipitants and different protein concentrations. Protein crystals were found in a monoclinic (C2) as well as an orthorhombic (P212121) space group. The crystals belonging to the monoclinic space group C2 had unit-cell parameters a = 103.4, b = 70.9, c = 54.9,Å, , = 109.3° for AsaP1_E294A, and a = 98.5, b = 74.5, c = 54.7,Å, , = 112.4° for AsaP1_E294Q. The unit-cell parameters of the orthorhombic crystal obtained for AsaP1_E294A were a = 57.9, b = 60.2, c = 183.6,Å. The crystals of the two different mutants diffracted X-rays beyond 2.0,Å resolution. [source] Crystallization and preliminary X-ray diffraction analysis of the , subunit Yke2 of the Gim complex from Saccharomyces cerevisiaeACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 6 2008Rebeca Pérez de Diego The Gim complex (GimC) from Saccharomyces cerevisiae is a heterohexameric protein complex, also known as prefoldin (PFD), which binds and stabilizes unfolded target polypeptides and subsequently delivers them to chaperonins for completion of folding. In this study, the crystallization and preliminary X-ray analysis of one of the , subunits of the Gim complex (Yke2) from S. cerevisiae are described. The purified protein was crystallized by the vapour-diffusion method, producing two types of crystals that belonged to the orthorhombic space group C222 or the primitive monoclinic space group P21. The unit-cell parameters for the C -centred orthorhombic crystal were a = 48.2, b = 168.86, c = 131.81,Å and the unit-cell parameters for the primitive monoclinic crystal were a = 47.83, b = 134.90, c = 81.50,Å, , = 100.71°. The Yke2 crystals diffracted to 4.2 and 3.1,Å resolution, respectively, on a rotating-anode generator under cryoconditions. This is the first report concerning the crystallization of a , subunit of a eukaryotic prefoldin. [source] Crystallization conditions and formation of orthorhombic paracetamol from ethanolic solutionJOURNAL OF PHARMACY AND PHARMACOLOGY: AN INTERNATI ONAL JOURNAL OF PHARMACEUTICAL SCIENCE, Issue 3 2002N. Al-Zoubi Orthorhombic paracetamol exhibits far better tabletability than the monoclinic form and its bulk crystallization from solution attracts much interest. In this study, temperature changes in supersaturated ethanolic solution have been recorded after seeding with orthorhombic crystals under different cooling temperatures (TC) and agitation rates (AR). Average cooling rate (CR), time for maximum temperature deviation (tmax) and area confined between curves of measured and reference temperature plots (AUC) were calculated and correlated with crystal yield (Y). The micromeritic (size and shape) and the compression properties, the density and the orthorhombic content of the crystalline product were evaluated and related to the main crystallization conditions applied (TC and AR). Conditions for optimal crystal yield and orthorhombic content were elucidated. It was found that crystal yield (Y) increased with AR and decreased with TC. The ratio tmax/CR provided good prediction of crystal yield (Y = 58.92 ,1.386 tmax/CR, r2 = 0.964 and P = 0.0001). TC and AR linearly affected crystal size and the size distribution, probably due to alterations in supersaturation, but they did not affect the crystal shape significantly. Density and compression properties (yield pressure and elastic recovery) were determined by the content of the orthorhombic form, which increased linearly with AR (P = 0.009) and with TC (P = 0.039) when agitation was between 300 and 500 rev min,1, while tmax decreased. At 700 rev min,1 orthorhombic content was maximized and became independent to TC. Higher orthorhombic content and crystal yield was expected for lower TC and for lower tmax, which corresponded to higher AR and might have also been affected by alteration of seeding and harvesting procedure. [source] 11-Methyl-2,3-benzodipyrrin-1-oneACTA CRYSTALLOGRAPHICA SECTION C, Issue 12 2004Raymond Bonnett The title compound {alternative names: 11-methyl-2,3-benzopyrromethenone and 3-[(1-methylpyrrol-2-yl)methylidene]-2,3-dihydro-1H -isoindol-1-one}, C14H12N2O, was prepared by the base-catalysed condensation of phthalimidine with 2-formyl-1-methylpyrrole; yellow orthorhombic crystals, space group Pbca, were obtained from ethanol. The molecule is almost planar, having Z(,)antiperiplanar geometry. The molecules are arranged in pairs with intermolecular hydrogen bonding between lactam functions. Comparison with literature values for polyalkyldipyrrin-1-ones shows that, apart from the local constraints of the benzene ring, the fused benzo ring has little effect on the molecular dimensions of the dipyrrin-1-one skeleton. [source] Pseudo-merohedral twinning and noncrystallographic symmetry in orthorhombic crystals of SIVmac239 Nef core domain bound to different-length TCR, fragmentsACTA CRYSTALLOGRAPHICA SECTION D, Issue 2 2010Walter M. Kim HIV/SIV Nef mediates many cellular processes through interactions with various cytoplasmic and membrane-associated host proteins, including the signalling , subunit of the T-cell receptor (TCR,). Here, the crystallization strategy, methods and refinement procedures used to solve the structures of the core domain of the SIVmac239 isolate of Nef (Nefcore) in complex with two different TCR, fragments are described. The structure of SIVmac239 Nefcore bound to the longer TCR, polypeptide (Leu51,Asp93) was determined to 3.7,Å resolution (Rwork = 28.7%) in the tetragonal space group P43212. The structure of SIVmac239 Nefcore in complex with the shorter TCR, polypeptide (Ala63,Arg80) was determined to 2.05,Å resolution (Rwork = 17.0%), but only after the detection of nearly perfect pseudo-merohedral crystal twinning and proper assignment of the orthorhombic space group P212121. The reduction in crystal space-group symmetry induced by the truncated TCR, polypeptide appears to be caused by the rearrangement of crystal-contact hydrogen-bonding networks and the substitution of crystallographic symmetry operations by similar noncrystallographic symmetry (NCS) operations. The combination of NCS rotations that were nearly parallel to the twin operation (k, h, ,l) and a and b unit-cell parameters that were nearly identical predisposed the P212121 crystal form to pseudo-merohedral twinning. [source] Imperfect pseudo-merohedral twinning in crystals of fungal fatty acid synthaseACTA CRYSTALLOGRAPHICA SECTION D, Issue 2 2009Simon Jenni The recent high-resolution structures of fungal fatty acid synthase (FAS) have provided new insights into the principles of fatty acid biosynthesis by large multifunctional enzymes. The crystallographic phase problem for the 2.6,MDa fungal FAS was initially solved to 5,Å resolution using two crystal forms from Thermomyces lanuginosus. Monoclinic crystals in space group P21 were obtained from orthorhombic crystals in space group P212121 by dehydration. Here, it is shown how this space-group transition induced imperfect pseudo-merohedral twinning in the monoclinic crystal, giving rise to a Moiré pattern-like interference of the two twin-related reciprocal lattices. The strategy for processing the twinned diffraction images and obtaining a quantitative analysis is presented. The twinning is also related to the packing of the molecules in the two crystal forms, which was derived from self-rotation function analysis and molecular-replacement solutions using a low-resolution electron microscopy map as a search model. [source] Differential effects of short affinity tags on the crystallization of Pyrococcus furiosus maltodextrin-binding proteinACTA CRYSTALLOGRAPHICA SECTION D, Issue 3 2002Matthew H. Bucher Pyrococcus furiosus maltodextrin-binding protein readily forms large orthorhombic crystals that diffract to high resolution. This protein was used as a model system to investigate the influence of five short affinity tags (His6, Arg5, Strep tag II, FLAG tag and the biotin acceptor peptide) on the formation of protein crystals and their ability to diffract X-rays. The results indicate that the amino-acid sequence of the tag can have a profound effect on both of these parameters. Consequently, the ability to obtain diffracting crystals of a particular protein may depend as much on which affinity tag is selected as it does on whether an affinity tag is used at all. [source] Intermonomer interactions in dimer of bovine heart cytochrome c oxidaseACTA CRYSTALLOGRAPHICA SECTION D, Issue 7 2001Soo Jae Lee The X-ray structure of bovine heart cytochrome c oxidase solved for orthorhombic crystals showed a dimeric structure stabilized by four subunit,subunit contacts, namely, subunit Vb,subunit Vb on the matrix side, subunit I,subunit VIa, subunit VIa,subunit I in the transmembrane region and subunit VIb,subunit VIb on the intermembrane side. The same intermonomer contacts as in the orthorhombic crystals were observed in both hexagonal and tetragonal crystals, the X-ray structures of which were determined by the molecular-replacement method. These results suggest that the dimeric structure also exists under physiological conditions. These contacts, especially the subunit IVa,subunit I contact, in which the N-terminal portion of subunit IVa is placed on the surface of subunit I near the dioxygen-reduction site, indicate that the function of the bovine heart enzyme is likely to be controlled by perturbation of the monomer,monomer association. [source] Preliminary crystallographic studies of an extremely thermostable KDG aldolase from Sulfolobus solfataricusACTA CRYSTALLOGRAPHICA SECTION D, Issue 11 2000Elaine J. Hendry Crystals have been grown of 2-keto-3-deoxygluconate aldolase (KDG aldolase) from the hyperthermophilic archaeon Sulfolobus solfataricus that diffract to 2.2,Å resolution. The enzyme catalyses the reversible aldol cleavage of 2-keto-3-dexoygluconate to pyruvate and glyceraldehyde, the third step of a modified non-phosphorylated Entner,Doudoroff pathway of glucose oxidation. S. solfataricus grows optimally at 353,K and the enzyme itself has a half-life of 2.5,h,at 373,K. Knowledge of the crystal structure of KDG aldolase will further understanding of the basis of protein hyperthermostability and create a target for site-directed mutagenesis of active-site residues, with the aim of altering substrate specificity. Three crystal forms have been obtained: orthorhombic crystals of space group P212121, which diffract to beyond 2.15,Å, monoclinic crystals of space group C2, which diffract to 2.2,Å, and cubic crystals of space group P4232, which diffract to 3.4,Å. [source] The 1.30,Å resolution structure of the Bacillus subtilis chorismate mutase catalytic homotrimerACTA CRYSTALLOGRAPHICA SECTION D, Issue 6 2000Jane E. Ladner The crystal structure of the Bacillus subtilis chorismate mutase, an enzyme of the aromatic amino acids biosynthetic pathway, was determined to 1.30,Å resolution. The structure of the homotrimer was determined by molecular replacement using orthorhombic crystals of space group P212121 with unit-cell parameters a = 52.2, b = 83.8, c = 86.0,Å. The ABC trimer of the monoclinic crystal structure [Chook et al. (1994), J. Mol. Biol.240, 476,500] was used as the starting model. The final coordinates are composed of three complete polypeptide chains of 127 amino-acid residues. In addition, there are nine sulfate ions, five glycerol molecules and 424 water molecules clearly visible in the structure. This structure was refined with aniosotropic temperature factors, has excellent geometry and a crystallographic R factor of 0.169 with an Rfree of 0.236. The three active sites of the macromolecule are at the subunit interfaces, with residues from two subunits contributing to each site. This orthorhombic crystal form was grown using ammonium sulfate as the precipitant; glycerol was used as a cryoprotectant during data collection. A glycerol molecule and sulfate ion in each of the active sites was found mimicking a transition-state analog. In this structure, the C-terminal tails of the subunits of the trimer are hydrogen bonded to residues of the active site of neighboring trimers in the crystal and thus cross-link the molecules in the crystal lattice. [source] Crystallization and preliminary X-ray studies of a galactose-specific lectin from the seeds of bitter gourd (Momordica charantia)ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 9 2010Thyageshwar Chandran A galactose-specific lectin from the seeds of bitter gourd (Momordica charantia) is a four-chain type II ribosome-inactivating protein (RIP) resulting from covalent association through a disulfide bridge between two identical copies of a two-chain unit. The available structural information on such four-chain RIPs is meagre. The bitter gourd lectin was therefore crystallized for structural investigation and the crystals have been characterized. It is anticipated that the structure of the orthorhombic crystals will be analysed using molecular replacement by taking advantage of its sequence, and presumably structural, homology to normal two-chain type II RIPs. [source] Expression, purification and preliminary X-ray diffraction analysis of the catalytic module of a ,-agarase from the flavobacterium Zobellia galactanivoransACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 4 2010Jan-Hendrik Hehemann Marine bacteria secrete specific glycoside hydrolases such as agarases to access polysaccharides from algal cell walls as a carbon and energy source. In an attempt to identify agarases with variable degradation patterns, a novel family GH16 ,-agarase from the marine bacterium Zobellia galactanivorans was expressed, purified and crystallized. The purified enzyme crystallized in two distinct forms that were grown by the hanging-drop vapour-diffusion method using polyethylene glycol as a precipitant. Hexagonal crystals belonging to space group P3121 diffracted to 2.2,Å resolution, whereas orthorhombic crystals belonging to space group P212121 diffracted to 1.5,Å resolution. [source] Crystallization and preliminary crystallographic analysis of cgHle, a homoserine acetyltransferase homologue, from Corynebacterium glutamicumACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 1 2009Christine Tölzer CgHle is an enzyme that is encoded by gene cg0961 from Corynebacterium glutamicum. The physiological function of cgHle is so far unclear. Bioinformatic annotations based on sequence homology indicated that cgHle may be an acetyl-CoA:homoserine acetyl transferase and as such may be involved in methionine biosynthesis, but recent evidence has shown that it is an esterase that catalyzes the hydrolysis of acetyl esters. Here, the crystallization of cgHle in two orthorhombic crystal forms, a trigonal crystal form and a monoclinic crystal form is described. The trigonal crystals have a solvent content of 83.7%, which is one of the highest solvent contents ever found for protein crystals. One of the orthorhombic crystals diffracted X-rays to at least 1.2,Å resolution. [source] Crystallization of the hydantoin transporter Mhp1 from Microbacterium liquefaciensACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 12 2008Tatsuro Shimamura The integral membrane protein Mhp1 from Microbacterium liquefaciens transports hydantoins and belongs to the nucleobase:cation symporter 1 family. Mhp1 was successfully purified and crystallized. Initial crystals were obtained using the hanging-drop vapour-diffusion method but diffracted poorly. Optimization of the crystallization conditions resulted in the generation of orthorhombic crystals (space group P212121, unit-cell parameters a = 79.7, b = 101.1, c = 113.8,Å). A complete data set has been collected from a single crystal to a resolution of 2.85,Å with 64,741 independent observations (94% complete) and an Rmerge of 0.12. Further experimental phasing methods are under way. [source] Expression, purification, crystallization and structure determination of two glutathione S -transferase-like proteins from Shewanella oneidensisACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 6 2008Bert Remmerie Genome analysis of Shewanella oneidensis, a Gram-negative bacterium with an unusual repertoire of respiratory and redox capabilities, revealed the presence of six glutathione S -transferase-like genes (sogst1,sogst6). Glutathione S -transferases (GSTs; EC 2.5.1.18) are found in all kingdoms of life and are involved in phase II detoxification processes by catalyzing the nucleophilic attack of reduced glutathione on diverse electrophilic substrates, thereby decreasing their reactivity. Structure,function studies of prokaryotic GST-like proteins are surprisingly underrepresented in the scientific literature when compared with eukaryotic GSTs. Here, the production and purification of recombinant SoGST3 (SO_1576) and SoGST6 (SO_4697), two of the six GST-like proteins in S. oneidensis, are reported and preliminary crystallographic studies of crystals of the recombinant enzymes are presented. SoGST3 was crystallized in two different crystal forms in the presence of GSH and DTT that diffracted to high resolution: a primitive trigonal form in space group P31 that exhibited merohedral twinning with a high twin fraction and a primitive monoclinic form in space group P21. SoGST6 yielded primitive orthorhombic crystals in space group P212121 from which diffraction data could be collected to medium resolution after application of cryo-annealing protocols. Crystal structures of both SoGST3 and SoGST6 have been determined based on marginal search models by maximum-likelihood molecular replacement as implemented in the program Phaser. [source] Structure of the T109S mutant of Escherichia coli dihydroorotase complexed with the inhibitor 5-fluoroorotate: catalytic activity is reflected by the crystal formACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 3 2007Mihwa Lee Crystals of a single-point mutant (T109S) of Escherichia coli dihydroorotase (DHOase) with diminished activity grown in the presence of l -dihydroorotate (l -DHO) are tetragonal, with a monomer in the asymmetric unit. These crystals are extremely unstable and disintegrate shortly after formation, which is followed by the growth of orthorhombic crystals from the remnants of the tetragonal crystals or at new nucleation sites. Orthorhombic crystals, for which a structure has previously been reported [Thoden et al. (2001), Biochemistry, 40, 6989,6997; Lee et al. (2005), J. Mol. Biol.348, 523,533], contain a dimer of DHOase in the asymmetric unit; the active site of one monomer contains the substrate N -carbamyl- l -aspartate (l -CA-asp) and the active site of the other monomer contains the product of the reaction, l -DHO. In the subunit with l -DHO in the active site, a surface loop (residues 105,115) is `open'. In the other subunit, with l -CA-asp in the active site, the loop folds inwards, forming specific hydrogen bonds from the loop to the l -CA-asp. The tetragonal crystal form can be stabilized by crystallization in the presence of the inhibitor 5-fluoroorotate (FOA), a product (l -DHO) mimic. Crystals of the complex of T109S DHOase with FOA are tetragonal, space group P41212, with unit-cell parameters a = b = 72.6, c = 176.1,Å. The structure has been refined to R and Rfree values of 0.218 and 0.257, despite severe anisotropy of the diffraction. In this structure, the flexible loops are both in the `open' conformation, which is consistent with FOA, like l -DHO, binding at both sites. The behaviour of the T109S mutant crystals of DHOase in the presence of l -DHO is explained by initial binding of l -DHO to both subunits, followed by slow conversion to l -CA-asp, with consequent movement of the flexible loop and dissolution of the crystals. Orthorhombic crystals are then able to grow in the presence of l -DHO and l -CA-asp. [source] | ||||||||||