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M Ammonium (m + ammonium)
Terms modified by M Ammonium Selected AbstractsESTABLISHMENT OF MINIMAL AND MAXIMAL TRANSCRIPT LEVELS FOR NITRATE TRANSPORTER GENES FOR DETECTING NITROGEN DEFICIENCY IN THE MARINE PHYTOPLANKTON ISOCHRYSIS GALBANA (PRYMNESIOPHYCEAE) AND THALASSIOSIRA PSEUDONANA (BACILLARIOPHYCEAE),JOURNAL OF PHYCOLOGY, Issue 4 2009Lee-Kuo Kang Nitrate transporter genes (Nrt2) encode high-affinity nitrate transporters in marine phytoplankton, and their transcript levels are potential markers of nitrogen deficiency in eukaryotic phytoplankton. For the proper interpretation of measured Nrt2 transcript abundances, a relative expression assay was proposed and tested in Isochrysis galbana Parke (Prymnesiophyceae) and Thalassiosira pseudonana (Hust.) Hasle et Heimdal (Bacillariophyceae). The minimal transcript levels of Nrt2 genes were achieved by the addition of 100 ,M ammonium, which led to a rapid decline in Nrt2 transcripts in 10,30 min. Experiments using a concentration series revealed that the effective dosage of ammonium to create a minimal transcript level of ,1 ,mol · mol,1 18S rRNA was ,25 ,M in both species. On the other hand, the addition of l -methionine sulfoximine (MSX), an inhibitor of glutamine synthetase, enhanced the Nrt2 transcript level in I. galbana but did not affect that in T. pseudonana. Nitrogen deprivation was used as an alternative means to create maximal Nrt2 transcript levels. By transferring cells into N-free medium for 24 h, Nrt2 transcript levels increased to ,90 ,mol · mol,1 18S rRNA in I. galbana, and to ,800 ,mol · mol,1 18S rRNA in T. pseudonana. The degree of nitrogen deficiency thus can be determined by comparing original Nrt2 transcript levels with the minimal and maximal levels. [source] The effect of a proline residue on the rate of growth and the space group of ,-spectrin SH3-domain crystalsACTA CRYSTALLOGRAPHICA SECTION D, Issue 12 2009Ana Cámara-Artigas ,-Spectrin SH3-domain (Spc-SH3) crystallization is characterized by very fast growth of the crystals in the presence of ammonium sulfate as a precipitant agent. The origin of this behaviour can be attributed to the presence of a proline residue that participates in a crystal contact mimicking the binding of proline-rich sequences to SH3 domains. This residue, Pro20, is located in the RT loop and is the main contact in one of the interfaces present in the orthorhombic Spc-SH3 crystal structures. In order to understand the molecular interactions that are responsible for the very fast crystal growth of the wild-type (WT) Spc-SH3 crystals, the crystal structure of a triple mutant in which the residues Ser19-Pro20-Arg21 in the RT loop have been replaced by Gly19-Asp20-Ser21 (GDS Spc-SH3 mutant) has been solved. The removal of the critical proline residue results in slower nucleation of the Spc-SH3 crystals and a different arrangement of the protein molecules in the unit cell, leading to a crystal that belongs to the tetragonal space group P41212, with unit-cell parameters a = b = 42.231, c = 93.655,Å, and that diffracts to 1.45,Å resolution. For both WT Spc-SH3 and the GDS mutant, light-scattering experiments showed that a dimer was formed in solution within a few minutes of the addition of 2,M ammonium sulfate at pH 6.5 and allowed the proposal of a mechanism for the nucleation and crystal growth of Spc-SH3 in which the Pro20 residue plays a key role in the rate of crystal growth. [source] Crystallization and preliminary X-ray analysis of cellobiose phosphorylase from Cellvibrio gilvusACTA CRYSTALLOGRAPHICA SECTION D, Issue 10 2004Masafumi Hidaka A recombinant cellobiose phosphorylase from Cellvibrio gilvus has been prepared and crystallized by the sitting-drop vapour-diffusion method using 10,mg,ml,1 purified enzyme, 1.5,M ammonium sulfate, 0.1,M MES buffer pH 7.0 and 5,mM glucose. A suitable crystal was obtained after 10,d incubation at 298,K. The crystal belongs to space group P21, with unit-cell parameters a = 84.77, b = 98.31, c = 104.04,Å, , = 102.73°. X-ray diffraction data to 2.1,Å resolution have been collected at KEK-PF BL-5A. [source] Crystallization and preliminary X-ray crystallographic studies on the fungal immunomodulatory protein Fve from the golden needle mushroom (Flammulina velutipes)ACTA CRYSTALLOGRAPHICA SECTION D, Issue 8 2003See Voon Seow The fungal immunomodulatory protein from the edible golden needle mushroom (Flammulina velutipes), designated Fve, is a single polypeptide consisting of 114 amino-acid residues. It is believed to trigger the mitogenic proliferation of T lymphocytes and Th1 cytokine production. Here, it is demonstrated that Fve forms a homodimer in nature. In order to understand the relationship between its structure and function, Fve was crystallized using the hanging-drop method; the protein formed well diffracting crystals within 3,5,d in 2.5% PEG 400, 2.0,M ammonium sulfate and 0.1,M Tris base buffer pH 8.5. The space group of the Fve crystals is either P43212 or P41212, with unit-cell parameters a = b = 96.92, c = 61.42,Å. The crystal contains two molecules per asymmetric unit and diffracts to 1.4,Å resolution when exposed to synchrotron radiation. [source] Crystallographic characterization of the PDZ1 domain of the human Na+/H+ exchanger regulatory factorACTA CRYSTALLOGRAPHICA SECTION D, Issue 5 2001Gordon Webster The Na+/H+ exchanger regulatory factor (NHERF) contains two PDZ domains that mediate the assembly of transmembrane and cytosolic proteins into functional signal transduction complexes. The human NHERF PDZ1 domain, which spans residues 11,99, interacts specifically with carboxy-terminal residues of the ,2 adrenergic receptor and the cystic fibrosis transmembrane conductance regulator. The NHERF PDZ1 was expressed in Escherichia coli as a soluble protein, purified and crystallized in the unbound form using the vapor-diffusion method with 2,M ammonium sulfate as the precipitant. Diffraction data were collected to 1.5,Å resolution using synchrotron radiation. The crystals belong to space group P3121 or P3221, with unit-cell parameters a = b = 51.6, c = 58.9,Å, and one molecule in the asymmetric unit. [source] Crystallization and preliminary structure determination of an intact human immunoglobulin, b12: an antibody that broadly neutralizes primary isolates of HIV-1ACTA CRYSTALLOGRAPHICA SECTION D, Issue 1 2001Erica Ollmann Saphire An intact human immunoglobulin with a full-length hinge has been crystallized for the first time in a form in which all of the Ig domains are ordered. The IgG1 antibody b12 is one of only three known monoclonal antibodies described that potently neutralize a broad range of HIV-1 primary isolates. It binds to an epitope overlapping the conserved CD4 binding site on the viral surface antigen gp120. Hexagonal crystals corresponding to space group R32 were grown from 0.8,M ammonium sulfate, with unit-cell parameters a = b = 271.3, c = 175.2,Å and one molecule per asymmetric unit. The crystals diffract to 2.8,Å and a preliminary molecular-replacement solution indicates that all 12 Ig domains of the antibody can be resolved. [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] Structure of the newly found green turtle egg-white ribonucleaseACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 7 2010Somporn Katekaew Marine green turtle (Chelonia mydas) egg-white ribonuclease (GTRNase) was crystallized from 1.1,M ammonium sulfate pH 5.5 and 30% glycerol using the sitting-drop vapour-diffusion method. The structure of GTRNase has been solved at 1.60,Å resolution by the molecular-replacement technique using a model based on the structure of RNase 5 (murine angiogenin) from Mus musculus (46% identity). The crystal belonged to the monoclinic space group C2, with unit-cell parameters a = 86.271, b = 34.174, c = 39.738,Å, , = 90, , = 102, , = 90°. GTRNase consists of three helices and seven ,-strands and displays the ,+, folding topology typical of a member of the RNase A superfamily. Superposition of the C, coordinates of GTRNase and RNase A superfamily members indicates that the overall structure is highly similar to that of angiogenin or RNase 5 from M. musculus (PDB code 2bwl) and RNase A from Bos taurus (PDB code 2blz), with root-mean-square deviations of 3.9 and 2.0,Å, respectively. The catalytic residues are conserved with respect to the RNase A superfamily. The three disulfide bridges observed in the reptilian enzymes are conserved in GTRNase, while one further disulfide bond is required for the structural stability of mammalian RNases. GTRNase is expressed in egg white and the fact that its sequence has the highest similarity to that of snapping turtle pancreatic RNase suggests that the GTRNase secreted from oviduct cells to form egg white is probably the product of the same gene as activated in pancreatic cells. [source] Crystallization and preliminary X-ray diffraction studies of hyperthermophilic archaeal Rieske-type ferredoxin (ARF) from Sulfolobus solfataricus P1ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 7 2010Asako Kounosu The hyperthermophilic archaeal Rieske-type [2Fe,2S] ferredoxin (ARF) from Sulfolobus solfataricus P1 contains a low-potential Rieske-type [2Fe,2S] cluster that has served as a tractable model for ligand-substitution studies on this protein family. Recombinant ARF harbouring a pET30a vector-derived N-terminal extension region plus a hexahistidine tag has been heterologously overproduced in Escherichia coli, purified and crystallized by the hanging-drop vapour-diffusion method using 0.05,M sodium acetate, 0.05,M HEPES, 2,M ammonium sulfate pH 5.5. The crystals diffracted to 1.85,Å resolution and belonged to the tetragonal space group P43212, with unit-cell parameters a = 60.72, c = 83.31,Å. The asymmetric unit contains one protein molecule. [source] Structure of photosynthetic glyceraldehyde-3-phosphate dehydrogenase (isoform A4) from Arabidopsis thaliana in complex with NADACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 6 2010Simona Fermani The crystal structure of the A4 isoform of photosynthetic glyceraldehyde-3-phosphate dehydrogenase (GAPDH) from Arabidopsis thaliana, expressed in recombinant form and complexed with NAD, is reported. The crystals, which were grown in 2.4,M ammonium sulfate and 0.1,M sodium citrate, belonged to space group I222. The asymmetric unit includes ten subunits, i.e. two independent tetramers plus a dimer that generates a third tetramer by a crystallographic symmetry operation. The crystal structure was solved by molecular replacement and refined to an R factor of 23.7% and an Rfree factor of 28.9% at 2.6,Å resolution. In the final model, each subunit binds one NAD+ molecule and two sulfates, which occupy the Ps and the Pi anion-binding sites. Detailed knowledge of this structure is instrumental for structural investigation of supramolecular complexes of A4 -GAPDH, phosphoribulokinase and CP12, which are involved in the regulation of photosynthesis in the model plant A. thaliana. [source] Crystallization and preliminary X-ray diffraction studies of the carbohydrate-recognition domain of SIGN-R1, a receptor for microbial polysaccharides and sialylated antibody on splenic marginal zone macrophagesACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 12 2009Noella Silva-Martin SIGN-R1, or CD209b, is a mouse C-type lectin receptor that is expressed at high levels on macrophages in lymphoid tissues, especially within the marginal zone of the spleen. SIGN-R1 can bind and mediate the uptake of various microbial polysaccharides, including dextrans, lipopolysaccharides and pneumococcal capsular polysaccharides. It has been shown that SIGN-R1 mediates the clearance of encapsulated pneumococcus, complement fixation via binding C1q independent of antibody and innate resistance to pneumococcal infection. Recently, SIGN-R1 has also been demonstrated to bind sialylated antibody and mediate its activity to suppress autoimmunity. The carbohydrate-recognition domain (CRD) of SIGN-R1 has been cloned and overexpressed in a soluble secretory form in mammalian Chinese hamster ovary (CHO) cells. The CRD protein of SIGN-R1 was purified from CHO cell-culture supernatant and concentrated for crystallization using the hanging-drop vapour-diffusion method at 291,K. Crystals grew from a mixture of 2,M ammonium sulfate in 0.1,M bis-tris pH 5.5. Single crystals, which belonged to the monoclinic space group C2 with unit-cell parameters a = 146.72, b = 92.77, c = 77.06,Å, , = 121.66°, allowed the collection of a full X-ray data set to a maximum resolution of 1.87,Å. [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] Expression, purification, crystallization and preliminary crystallographic analysis of an endo-1,5-,- l -arabinanase from hyperthermophilic Thermotoga petrophilaACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 9 2009Fabio Marcio Squina The endo-1,5-,- l -arabinanases belonging to glycoside hydrolase family 43 are of great industrial interest for use in food technology, organic synthesis and biofuel production owing to their ability to catalyze the hydrolysis of ,-1,5-arabinofuranosidic bonds in arabinose-containing polysaccharides. In this work, Thermotoga petrophila endo-1,5-,- l -arabinanase, a GH43-family member, has been cloned, overexpressed, purified and crystallized. Single crystals were obtained from a solution containing 0.1,M MES buffer pH 6.5, 0.8,M ammonium sulfate, 0.1,M EDTA, 0.1,Ml -proline and 5%(v/v) dioxane. X-ray diffraction data were collected to a resolution of 2.86,Å using synchrotron radiation and the diffraction pattern was indexed in the tetragonal space group P422, with unit-cell parameters a = b = 83.71, c = 408.25,Å. [source] Purification and preliminary X-ray crystallographic studies of ,-microseminoprotein from human seminal plasmaACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 5 2009Vijay Kumar ,-Microseminoprotein (,-MSP) is a small cysteine-rich protein with a molecular mass of 10,kDa. It was first isolated from human seminal plasma and has subsequently been identified from several species. Comparison of the amino-acid sequences of ,-MSP proteins suggests that the protein is a rapidly evolving protein. The function of ,-MSP is poorly understood. Furthermore, no crystal structure has been reported of any ,-MSP; therefore, determination of the crystal structure of ,-MSP is the foremost task in order to understand the function of this protein completely. Here, the purification, crystallization and preliminary X-ray diffraction analysis of ,-MSP from human seminal plasma are described. The protein was purified using anion-exchange and size-exclusion chromatography and the purified protein was crystallized using 0.1,M ammonium sulfate, 0.1,M HEPES buffer pH 7.0 and 20%(w/v) PEG 3350. The crystals belonged to the tetragonal space group P4322 and contained three ,-MSP molecules in the asymmetric unit. X-ray intensity data were collected to 2.4,Å resolution. [source] Purification, crystallization and preliminary crystallographic analysis of deoxyuridine triphosphate nucleotidohydrolase from Arabidopsis thalianaACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 5 2007Mamta Bajaj The deoxyuridine triphosphate nucleotidohydrolase gene from Arabidopsis thaliana was expressed and the gene product was purified. Crystallization was performed by the hanging-drop vapour-diffusion method at 298,K using 2,M ammonium sulfate as the precipitant. X-ray diffraction data were collected to 2.2,Å resolution using Cu,K, radiation. The crystal belongs to the orthorhombic space group P212121, with unit-cell parameters a = 69.90, b = 70.86,Å, c = 75.55,Å. Assuming the presence of a trimer in the asymmetric unit, the solvent content was 30%, with a VM of 1.8,Å3,Da,1. [source] Overexpression, purification and crystallization of tyrosyl-tRNA synthetase from the hyperthermophilic archaeon Aeropyrum pernix K1ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 11 2005Jun Iwaki Hyperthermophilic archaeal tyrosyl-tRNA synthetase from Aeropyrum pernix K1 was cloned and overexpressed in Escherichia coli. The expressed protein was purified by Cibacron Blue affinity chromatography following heat treatment at 363,K. Crystals suitable for X-ray diffraction studies were obtained under optimized crystallization conditions in the presence of 1.5,M ammonium sulfate using the hanging-drop vapour-diffusion method. The crystals belonged to the tetragonal space group P43212, with unit-cell parameters a = b = 66.1, c = 196.2,Å, and diffracted to beyond 2.15,Å resolution at 100,K. [source] Crystallization and preliminary X-ray crystallographic analysis of agkicetin-C from Deinagkistrodon acutus venomACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 1 2005Gufeng Xu The crystallization and preliminary crystallographic analysis of agkicetin-C, a well known platelet glycoprotein Ib (GPIb) antagonist from the venom of Deinagkistrodon acutus found in Anhui Province, China is reported. Crystals of agkicetin-C suitable for structure determination were obtained from 1.8,M ammonium sulfate, 40,mM MES pH 6.5 with 2%(v/v) PEG 400. Interestingly, low buffer concentrations of MES seem to be necessary for crystal growth. The crystals of agkicetin-C belong to space group C2, with unit-cell parameters a = 177.5, b = 97.7, c = 106.8,Å, , = 118.5°, and diffract to 2.4,Å resolution. Solution of the phase problem by the molecular-replacement method shows that there are four agkicetin-C molecules in the asymmetric unit, with a VM value of 3.4,Å3,Da,1, which corresponds to a high solvent content of approximately 64%. Self-rotation function calculations show a single well defined non-crystallographic twofold axis with features that may represent additional elements of non-crystallographic symmetry. [source] | ||||||||||