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Preliminary X-ray Diffraction Analysis (preliminary + x-ray_diffraction_analysis)

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Selected Abstracts

Crystallization and preliminary X-ray diffraction analysis of a [2Fe,2S] ferredoxin (FdVI) from Rhodobacter capsulatus

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 2 2001
Jean Armengaud
A [2Fe,2S] ferredoxin found in the photosynthetic bacterium Rhodobacter capsulatus has been purified in recombinant form from Escherichia coli. This protein, called FdVI, resembles ferredoxins involved in iron,sulfur cluster biosynthesis in various prokaryotic and eukaryotic cells. Purified recombinant FdVI was recovered in high yields and appeared to be indistinguishable from the genuine R. capsulatus ferredoxin based on UV,visible absorption and EPR spectroscopy and mass spectrometry. FdVI has been crystallized in the oxidized state by a sitting-drop vapour-diffusion technique using sodium formate as precipitant. Seeding larger drops from a previous hanging-drop-grown small crystal resulted in the formation of long red,brown prismatic needles. Preliminary X-ray diffraction analysis indicated that FdVI crystals are orthorhombic and belong to the space group P212121, with unit-cell parameters a = 45.87, b = 49.83, c = 54.29 Å. [source]

Crystallization and preliminary X-ray diffraction analysis of rat autotaxin

ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 9 2010
Jacqueline E. Day
Rat autotaxin has been cloned, expressed, purified to homogeneity and crystallized via hanging-drop vapour diffusion using PEG 3350 as precipitant and ammonium iodide and sodium thiocyanate as salts. The crystals diffracted to a maximum resolution of 2.05,Å and belonged to space group P1, with unit-cell parameters a = 53.8, b = 63.3, c = 70.5,Å, , = 98.8, , = 106.2, , = 99.8°. Preliminary X-ray diffraction analysis indicated the presence of one molecule per asymmetric unit, with a solvent content of 47%. [source]

Crystallization and preliminary X-ray diffraction analysis of the complex of Kunitz-type tamarind trypsin inhibitor and porcine pancreatic trypsin

ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 11 2009
Sakshi Tomar
The complex of Tamarindus indica Kunitz-type trypsin inhibitor and porcine trypsin has been crystallized by the sitting-drop vapour-diffusion method using ammonium acetate as precipitant and sodium acetate as buffer. The homogeneity of complex formation was checked by size-exclusion chromatography and further confirmed by reducing SDS,PAGE. The crystals diffracted to 2.0,Å resolution and belonged to the tetragonal space group P41, with unit-cell parameters a = b = 57.1, c = 120.1,Å. Preliminary X-ray diffraction analysis indicated the presence of one unit of inhibitor,trypsin complex per asymmetric unit, with a solvent content of 45%. [source]

Expression, purification, crystallization and preliminary X-ray analysis of the N-terminal domain of GNBP3 from Drosophila melanogaster

ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 9 2009
Yumiko Mishima
Gram-negative bacteria-binding protein 3 (GNBP3) is a pattern-recognition receptor which contributes to the defensive response against fungal infection in Drosophila. The protein consists of an N-terminal domain, which is considered to recognize ,-glucans from the fungal cell wall, and a C-terminal domain, which is homologous to bacterial glucanases but devoid of activity. The N-terminal domain of GNBP3 (GNBP3-Nter) was successfully purified after expression in Drosophila S2 cells. Diffraction-quality crystals were produced by the hanging-drop vapour-diffusion method using PEG 2000 and PEG 8000 as precipitants. Preliminary X-ray diffraction analysis revealed that the GNBP3-Nter crystals belonged to the monoclinic space group C2, with unit-cell parameters a = 134.79, b = 30.55, c = 51.73,Å, , = 107.4°, and diffracted to 1.7,Å using synchrotron radiation. The asymmetric unit is expected to contain two copies of GNBP3-Nter. Heavy-atom derivative data were collected and a samarium derivative showed one high-occupancy site per molecule. [source]

Crystallization and preliminary crystallographic characterization of the extrinsic PsbP protein of photosystem II from Spinacia oleracea

ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 2 2009
J. Kohoutová
Preliminary X-ray diffraction analysis of the extrinsic PsbP protein of photosystem II from spinach (Spinacia oleracea) was performed using N-terminally His-tagged recombinant PsbP protein overexpressed in Escherichia coli. Recombinant PsbP protein (thrombin-digested recombinant His-tagged PsbP) stored in bis-Tris buffer pH 6.00 was crystallized using the sitting-drop vapour-diffusion technique with PEG 550 MME as a precipitant and zinc sulfate as an additive. SDS,PAGE analysis of a dissolved crystal showed that the crystals did not contain the degradation products of recombinant PsbP protein. PsbP crystals diffracted to 2.06,Å resolution in space group P212121, with unit-cell parameters a = 38.68, b = 46.73, c = 88.9,Å. [source]

Preliminary X-ray diffraction analysis of the cytoplasmic N-terminal domain of the Na/HCO3 cotransporter NBCe1-A

ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 6 2006
Harindarpal S. Gill
The N-terminal cytoplasmic domain of the Na+ -coupled HCO cotransporter NBCe1-A (NtNBCe1) has been linked with proximal renal tubular acidosis. In a previous purification study of recombinant NtNBCe1, crystal growth at a suboptimal protein concentration (<1,mg,ml,1) yielded small single diamond-shaped crystals that diffracted poorly. In the present study, by increasing the protein concentration 50-fold, the crystal size was doubled and robustness was also improved. Crystal annealing made the crystals suitable for X-ray diffraction. The crystals either belong to space group P3121 or P31 with pseudo P3121 symmetry, with unit-cell parameters a = 51.7, b = 51.7, c = 200.6,Å, , = , = 90, , = 120°, and diffract X-rays to 3.0,Å resolution. The calculated Matthews number is 1.9,Å3,Da,1, with two monomers of molecular weight ,83,kDa in the asymmetric unit. The molecular- replacement packing solution shows that the molecules form dimers by a domain-swapping mechanism. [source]

Crystallization and preliminary X-ray diffraction analyses of the homodimeric glycine decarboxylase (P-protein) from the cyanobacterium Synechocystis sp.

ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 2 2010
PCC 680
Glycine decarboxylase, or P-protein, is a major enzyme that is involved in the C1 metabolism of all organisms and in the photorespiratory pathway of plants and cyanobacteria. The protein from Synechocystis sp. PCC 6803 is a homodimer with a mass of 215,kDa. Recombinant glycine decarboxylase was expressed in Escherichia coli and purified by metal-affinity, ion-exchange and gel-filtration chromatography. Crystals of P-protein that diffracted to a resolution of 2.1,Å were obtained using the hanging-drop vapour-diffusion method at 291,K. X-ray diffraction data were collected from cryocooled crystals using synchrotron radiation. The crystals belonged to space group P212121, with unit-cell parameters a = 96.30, b = 135.81, c = 179.08,Å. [source]

Cloning, expression, purification and crystallization as well as X-ray fluorescence and preliminary X-ray diffraction analyses of human ADP-ribosylhydrolase 1

ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 5 2009
Stefan Kernstock
Human ADP-ribosylhydrolase 1 (hARH1, ADPRH) cleaves the glycosidic bond of ADP-ribose attached to an Arg residue of a protein. hARH1 has been cloned, expressed heterologously in Escherichia coli, purified and crystallized in complex with K+ and ADP. The orthorhombic crystals contained one monomer per asymmetric unit, exhibited a solvent content of 43% and diffracted X-rays to a resolution of 1.9,Å. A prerequisite for obtaining well diffracting crystals was the performance of X-ray fluorescence analysis on poorly diffracting apo hARH1 crystals, which revealed the presence of trace amounts of K+ in the crystal. Adding K-ADP to the crystallization cocktail then resulted in a crystal of different morphology and with dramatically improved diffraction properties. [source]

Crystallization and preliminary X-ray diffraction analyses of several forms of the CfaB major subunit of enterotoxigenic Escherichia coli CFA/I fimbriae

ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 3 2009
Yong-Fu Li
Enterotoxigenic Escherichia coli (ETEC), a major global cause of diarrhea, initiates the pathogenic process via fimbriae-mediated attachment to the small intestinal epithelium. A common prototypic ETEC fimbria, colonization factor antigen I (CFA/I), consists of a tip-localized minor adhesive subunit CfaE and the stalk-forming major subunit CfaB, both of which are necessary for fimbrial assembly. To elucidate the structure of CFA/I at atomic resolution, three recombinant proteins were generated consisting of fusions of the minor and major subunits (CfaEB) and of two (CfaBB) and three (CfaBBB) repeats of the major subunit. Crystals of CfaEB diffracted X-rays to 2.1,Å resolution and displayed the symmetry of space group P21. CfaBB exhibited a crystal diffraction limit of 2.3,Å resolution and had the symmetry of space group P21212. CfaBBB crystallized in the monoclinic space group C2 and diffracted X-rays to 2.3,Å resolution. These structures were determined using the molecular-replacement method. [source]

Crystallization and preliminary X-ray diffraction analysis of homing endonuclease I- Tsp061I

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 11 2004
Takahito Imagawa
Two crystal forms, rhombohedral and hexagonal, of a homing endonuclease from Thermoproteus sp. IC-061 (I- Tsp0611) were obtained by the hanging-drop and sitting-drop method, respectively. The hexagonal crystals belong to space group P6322, with unit-cell parameters a = b = 111.4, c = 97.6,Å, and diffract to 3.2,Å resolution on beamline BL44 at SPring-8 (Harima, Japan). The rhombohedral crystals belong to space group R32, with unit-cell parameters a = b = 95.4, c = 192.9,Å, and diffract to 2.7,Å resolution using a Cu,K, rotating-anode generator with an R-AXIS VII detector. The crystal asymmetric unit contained one protein molecule and the solvent contents of the two crystal forms were estimated to be 68.3 and 67.6% by volume, respectively. [source]

Purification, crystallization and preliminary X-ray diffraction analysis of human oncoprotein SET/TAF-1,

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 4 2004
Shinsuke Muto
The human oncoprotein SET/TAF-1, has been crystallized by the sitting-drop vapour-diffusion method using ammonium sulfate as a precipitant. The crystal belongs to space group C2, with unit-cell parameters a = 119.6, b = 62.8, c = 61.0,Å, , = 89.7°, and contains two molecules in the asymmetric unit. A complete data set was collected to 2.8,Å resolution using synchrotron radiation. [source]

Crystallization and preliminary X-ray diffraction analysis of KsgA, a universally conserved RNA adenine dimethyltransferase in Escherichia coli

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 8 2003
Heather C. O'Farrell
The bacterial enzyme KsgA catalyzes the transfer of a total of four methyl groups from S -adenosylmethionine (SAM) to two adjacent adenosines in 16S rRNA. These modified adenosines are universally conserved in all species of eubacteria, eukaryotes and archaebacteria studied. Recombinant KsgA from Escherichia coli was overexpressed as a His-tagged fusion protein and purified. The recombinant protein was crystallized using PEG 4000 as a precipitant. The crystals belong to space group C2 and diffract X-rays to a resolution of 1.9,Å. The unit-cell parameters are a = 173.9, b = 38.4, c = 83.0,Å, , = 90.0°. Structure determination using the molecular-replacement method is at the early stages of refinement. [source]

Crystallization and preliminary X-ray diffraction analysis of thermophilic imidase from pig liver

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 5 2003
Cheng-Yang Huang
Imidase is an enzyme, also known as dihydropyrimidinase (EC 3.5.2.2), hydantoinase, dihydropyrimidine hydrase or dihydropyrimidine amidohydrolase, that catalyzes the reversible hydrolysis of 5,6-­dihydrouracil to 3-ureidopropionate and many other imides. Substrate specificity, metal content and amino-acid sequence all differ significantly between bacterial and mammalian imide-hydrolyzing enzymes. In this study, a thermophilic imidase was isolated from pig liver and crystallized. Two kinds of imidase crystals were grown by the hanging-drop vapour-diffusion method using polyethylene glycol MME 5000 and 2-propanol as precipitants. One belongs to the triclinic P1 space group, with unit-cell parameters a = 96.35, b = 96.87, c = 154.87,Å, , = 82.10, , = 72.54, , = 77.19°, and the other belongs to the orthorhombic C2221 space group, with unit-cell parameters a = 113.92, b = 157.22, c = 156.21,Å. [source]

Cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of HP1352, a putative DNA methyltransferase in Helicobacter pylori

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 4 2003
Katja Schirwitz
The putative methyltransferase gene HP1352 from Helicobacter pylori strain 26695 was heterologously expressed in Escherichia coli. The 359-amino-acid gene product was purified and crystallized. The crystals belong to space group I212121 and show diffraction to at least 2.5,Å resolution. The unit-cell parameters are a = 69.6, b = 86.6, c = 140.0,Å. A greater than 90% complete native data set has been collected and structure determination using the molecular-replacement method is ongoing. [source]

Crystallization and preliminary X-ray diffraction analysis of recombinant hydrolase domain of 10-­formyltetrahydrofolate dehydrogenase

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 10-2 2002
Alexander A. Chumanevich
10-Formyltetrahydrofolate dehydrogenase (FDH) is an abundant enzyme in liver cytosol. It is important for the regulation of 10-­formyltetrahydrofolate/tetrahydrofolate pools, for de novo purine biosynthesis and for the removal of formate in the form of CO2. The enzyme is a natural fusion of two unrelated genes and consists of two functional catalytic domains. Here, the crystallization of the N-­terminal domain of FDH is reported. This domain binds folate and functions as a 10-formyltetrahydrofolate hydrolase. The crystals grow as either spear-shaped needles or large plates, with the largest crystals reaching dimensions of 1.2 × 0.2 × 0.05,mm. Diffraction analysis revealed the space group to be P21212, with unit-cell parameters a = 100.00, b = 64.63, c = 64.59,Å. Based on the estimated solvent content, there is one 34,kDa molecule in the asymmetric unit. A native data set extending to 2.3,Å resolution has been collected with good merging statistics. [source]

Purification, crystallization and preliminary X-ray diffraction analysis of yeast nucleosome-assembly factor Cia1p

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 10-2 2002
Balasundaram Padmanabhan
Yeast Cia1p is a homologue of human CIA (CCG1-interacting factor A), which possesses nucleosome-assembly activity and interacts with the human TFIID subunit CCG1 and the C-terminal domain of histone H3. The yeast Cia1p without the C-terminal polyanionic stretch has been expressed in Escherichia coli, purified to homogeneity and crystallized by the hanging-drop vapour-diffusion method using PEG 8000 as precipitant. The protein was crystallized in orthorhombic space group P212121, with unit-cell parameters a = 106.70, b = 46.92, c = 40.60,Å and one molecule in the asymmetric unit. The crystal diffracted beyond 2.95,Å resolution using synchrotron radiation. [source]

Crystallization and preliminary X-ray diffraction analysis of glutamate dehydrogenase from an aerobic hyperthermophilic archaeon, Aeropyrum pernix K1

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 8 2002
Mohammad W. Bhuiya
Glutamate dehydrogenase from an aerobic hyperthermophilic archaeon, Aeropyrum pernix K1, was crystallized by the hanging-drop vapour-diffusion method using polyethylene glycol (PEG) 400 as the precipitant. The crystals belong to the hexagonal space group P63, with unit-cell parameters a = b = 98.9, c = 394.8,Å, , = , = 90, , = 120°. The asymmetric unit contained one hexamer of the enzyme, giving a crystal volume per enzyme mass (VM) of 1.98,Å3,Da,1 and a solvent content of 37.3%. The X-ray diffraction data were collected to a resolution of 3.0,Å at the BL6B beamline in the Photon Factory with an overall Rsym of 13.8% and a completeness of 87.1%. [source]

Crystallization and preliminary X-ray diffraction analysis of shikimate kinase from Mycobacterium tuberculosis in complex with MgADP

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 12 2001
Yijun Gu
Shikimate kinase (SK) from Mycobacterium tuberculosis (Mt) was overexpressed in Escherichia coli, purified and cocrystallized with MgADP in hanging drops using the vapor-diffusion procedure with PEG 4000 and 2-propanol as precipitants at pH 7.5. The crystal of MtSK,MgADP, which diffracted to 2.2,Å resolution, belonged to space group P3221 or P3121, with unit-cell parameters a = b = 64.01, c = 92.41,Å. There was one MtSK molecule in the asymmetric unit. Molecular-replacement trials with the crystal structure of SK from Erwinia chrysanthemi (PDB code 1shk) and adenylate kinase (PDB code 1ake) as search models were not successful. Heavy-atom derivative screening is in progress. [source]

Crystallization and preliminary X-ray diffraction analysis of cytotoxic ribonucleases from bullfrog Rana catesbeiana

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 11 2001
Jyung-Hurng Liu
RC-RNases are ribonucleases from Rana catesbeiana oocytes with pyrimidine,guanine sequence specificity. They also possess cell cytotoxicity and lectin activity. Protein crystals of three RC-RNase isozymes, RC-RNase 3, RC-RNase 4 and RC-RNase 6, were grown in various crystal systems under different conditions. Crystals of RC-­RNase3 belong to the orthorhombic C2221 space group, with unit-cell parameters a = 66.66, b = 97.38, c = 85.74,Å. Crystals of RC-­RNase 4 belong to the trigonal space group P31 or P32, with unit-cell parameters a = b = 32.22, c = 92.12,Å. Crystals of RC-RNase 6 complexed with cytidylyl 2,-5, guanosine belong to the tetragonal space group P41212 or P43212, with unit-cell parameters a = b = 61.80, c = 65.96,Å. [source]

Crystallization and preliminary X-ray diffraction analysis of a [2Fe,2S] ferredoxin (FdVI) from Rhodobacter capsulatus

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 2 2001
Jean Armengaud
A [2Fe,2S] ferredoxin found in the photosynthetic bacterium Rhodobacter capsulatus has been purified in recombinant form from Escherichia coli. This protein, called FdVI, resembles ferredoxins involved in iron,sulfur cluster biosynthesis in various prokaryotic and eukaryotic cells. Purified recombinant FdVI was recovered in high yields and appeared to be indistinguishable from the genuine R. capsulatus ferredoxin based on UV,visible absorption and EPR spectroscopy and mass spectrometry. FdVI has been crystallized in the oxidized state by a sitting-drop vapour-diffusion technique using sodium formate as precipitant. Seeding larger drops from a previous hanging-drop-grown small crystal resulted in the formation of long red,brown prismatic needles. Preliminary X-ray diffraction analysis indicated that FdVI crystals are orthorhombic and belong to the space group P212121, with unit-cell parameters a = 45.87, b = 49.83, c = 54.29 Å. [source]

Crystallization and preliminary X-ray diffraction analysis of protein l -isoaspartyl O -­methyltransferase from wheat germ

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 2 2001
Michelle D. Amaral
Wheat-germ protein l -isoaspartyl O -methyltransferase (WPIMT) can initiate the conversion of l -isoaspartyl residues in a protein or peptide, which accumulate during the aging process in wheat-germ seeds, to normal l -aspartyl groups. The recombinant protein of WPIMT was overexpressed in Escherichia coli and purified to homogeneity. The protein was crystallized in the presence of S -­adenosine- l -homocysteine using 2-methyl-2,4-pentanediol. Preliminary X-ray analysis indicated a tetragonal space group P41212 or P43212, with unit-cell parameters a = b = 77.3, c = 152.9,Å for cryofrozen crystals at 90,K. The crystals diffracted to 3.3,Å and contain two molecules per asymmetric unit. [source]

Crystallization and preliminary X-ray diffraction analysis of a eumenine mastoparan toxin: a new class of mast-cell degranulating peptide in the wasp venom

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 11 2000
F. Canduri
Mastoparans are tetradecapeptides found to be the major component of vespid venoms. A mastoparan toxin isolated from the venom of Anterhynchium flavomarginatum micado has been crystallized and X-­ray diffraction data collected to 2.7,Å resolution using a synchrotron-radiation source. Crystals were determined to belong to the space group P6222 (P6422). This is the first mastoparan to be crystallized and will provide further insights into the conformational significance of mastoparan toxins with respect to their potency and activity in G-protein regulation. [source]

Crystallization and preliminary X-ray diffraction analysis of a recombinant cysteine-free mutant of crmA

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 11 2000
Miljan Simonovic
CrmA is an unusual serpin that has a reactive-center loop one residue shorter than other members of the superfamily. Most interestingly, crmA has inhibitory activity against both cysteine and serine proteinases involved in the regulation of cell apoptosis. The three-dimensional structure of crmA will give insight into the mechanism that this serpin employs to inhibit both cysteine and the serine proteinases, as well as help to explain the significance of the shorter reactive-center loop. The monodisperse cysteine-free mutant of crmA was crystallized in the presence of phosphate salts. Crystals diffract to 2.90,Å and belong to space group P212121, with unit-cell parameters a = 42.67, b = 93.15, c = 101.63,Å. [source]

Crystallization and preliminary X-ray diffraction analysis of a rat biliverdin reductase

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 9 2000
Danyu Sun
Biliverdin reductase (BVR) catalyzes the final step of haem degradation and converts biliverdin to bilirubin using NAD(P)H as an electron donor. This paper deals with the first crystallization and preliminary crystallographic study of recombinant rat BVR expressed in Escherichia coli. Crystals of BVR were obtained by the sitting-drop vapour-diffusion method. Using synchrotron radiation at station BL44B2 of SPring-8, Japan, BVR diffraction data were collected to 1.6,Å resolution. Crystals belong to the orthorhombic space group P212121, with unit-cell parameters a = 58.89, b = 70.41, c = 87.76,Å. The complete determination of the crystallographic structure is currently in progress using MAD (multiwavelength anomalous diffraction) data from an Ir-derivative crystal. [source]

Crystallization and preliminary X-ray diffraction analysis of Thermus thermophilus prolyl-tRNA synthetase

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 2 2000
Anna Yaremchuk
Prolyl-tRNA synthetase from Thermus thermophilus (ProRSTT) was purified to homogeneity using a five-step purification procedure and was crystallized using ethylene glycol as a precipitant. Crystals of ProRSTT belong to the space group P21212, with unit-cell parameters a = 132, b = 191, c = 125,Å, have two homodimers per asymmetric unit and diffract to 2.4,Å resolution. A complete native data set to 2.43,Å resolution has been collected and a data set from ProRSTT in complex with proline has been collected to 2.9,Å resolution. [source]

Crystallization and preliminary X-ray diffraction analysis of rat autotaxin

ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 9 2010
Jacqueline E. Day
Rat autotaxin has been cloned, expressed, purified to homogeneity and crystallized via hanging-drop vapour diffusion using PEG 3350 as precipitant and ammonium iodide and sodium thiocyanate as salts. The crystals diffracted to a maximum resolution of 2.05,Å and belonged to space group P1, with unit-cell parameters a = 53.8, b = 63.3, c = 70.5,Å, , = 98.8, , = 106.2, , = 99.8°. Preliminary X-ray diffraction analysis indicated the presence of one molecule per asymmetric unit, with a solvent content of 47%. [source]

Cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of SAICAR synthase from Streptococcus suis serotype 2

ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 8 2010
Xia Cheng
Phosphoribosylaminoimidazole-succinocarboxamide synthase (SAICAR synthase) plays an essential role in the de novo biosynthesis of purine nucleotides. In this study, the SAICAR synthase from Streptococcus suis was cloned and overexpressed in Escherichia coli. The subsequent product was purified and crystallized using the hanging-drop vapour-diffusion method. The crystals diffracted to 2.8,Å resolution and belonged to space group P2, with unit-cell parameters a = 70.2, b = 52.2, c = 153.9,Å, , = 102.8°. [source]

Expression, crystallization and preliminary X-ray diffraction analysis of the CMM2 region of the Arabidopsis thaliana Morpheus' molecule 1 protein

ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 8 2010
Tom J. Petty
Of the known epigenetic control regulators found in plants, the Morpheus' molecule 1 (MOM1) protein is atypical in that the deletion of MOM1 does not affect the level of epigenetic marks controlling the transcriptional status of the genome. A short 197-amino-acid fragment of the MOM1 protein sequence can complement MOM1 deletion when coupled to a nuclear localization signal, suggesting that this region contains a functional domain that compensates for the loss of the full-length protein. Numerous constructs centred on the highly conserved MOM1 motif 2 (CMM2) present in these 197 residues have been generated and expressed in Escherichia coli. Following purification and crystallization screening, diamond-shaped single crystals were obtained that diffracted to ,3.2,Å resolution. They belonged to the trigonal space group P3121 (or P3221), with unit-cell parameters a = 85.64, c = 292.74,Å. Structure determination is ongoing. [source]

Crystallization and preliminary X-ray diffraction analysis of the putative aldose 1-epimerase YeaD from Escherichia coli

ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 8 2010
Weijie You
Escherichia coli YeaD (ecYeaD) is suggested to be a member of the galactose mutarotase-like superfamily. Galactose mutarotase is an enzyme that converts ,-galactose to ,-galactose. The known structures of these galactose mutarotase-like proteins are similar to those of galactose mutarotases, with the catalytic residues being conserved, but there are some differences between them in the substrate-binding pocket. In order to reveal the specificity of ecYeaD, a three-dimensional structure is essential. Full-length ecYeaD with an additional 6×His tag at the C-terminus was crystallized by the hanging-drop vapour-diffusion method using polyethylene glycol 4000 as a precipitant at 283,K. An X-ray diffraction data set was collected to a resolution of 1.9,Å from a single flash-cooled crystal that belonged to space group P212121. [source]

Crystallization and preliminary X-ray diffraction analysis of Pseudomonas aeruginosa phosphorylcholine phosphatase

ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 8 2010
Lisandro H. Otero
Pseudomonas aeruginosa phosphorylcholine phosphatase (PchP) catalyzes the hydrolysis of phosphorylcholine to produce choline and inorganic phosphate. Phosphorylcholine is released by the action of haemolytic phospholipase C (PlcH) on phosphatidylcholine or sphingomyelin. PchP belongs to the HAD superfamily and its activity is dependent on Mg2+, Zn2+ or Cu2+. The possible importance of PchP in the pathogenesis of P. aeruginosa, the lack of information about its structure and its low identity to other members of this family led us to attempt its crystallization in order to solve its three-dimensional structure. Crystals of the protein have been grown and diffraction data have been obtained to 2.7,Å resolution. The crystals belonged to the monoclinic space group C2, with unit-cell parameters a = 137.16, b = 159.15, c = 73.31,Å, , = 117.89°. Statistical analysis of the unit-cell contents and the self-rotation function suggest a tetrameric state of the molecule with 222 point-group symmetry. [source]