scholarly journals Purification, crystallization and preliminary crystallographic analysis of histone lysine demethylase NO66 fromHomo sapiens

2012 ◽  
Vol 68 (7) ◽  
pp. 764-766 ◽  
Author(s):  
Xing Zhou ◽  
Yue Tao ◽  
Minhao Wu ◽  
Dandan Zhang ◽  
Jianye Zang
Keyword(s):  
Crystallographic Analysis ◽  
Diffusion Method ◽  
Amino Acid Residues ◽  
X Ray Diffraction ◽  
Cell Parameters ◽  
Histone Lysine Demethylase ◽  
Jmjc Domain ◽  
Lysine Demethylase

NO66 is a JmjC domain-containing histone demethylase with specificity towards histone H3 methylated on both Lys4 and Lys36in vitroandin vivo. A fragment of NO66 lacking the N-terminal 167 amino-acid residues was overexpressed inEscherichia coli, purified and crystallized using the sitting-drop vapour-diffusion method. X-ray diffraction data were collected to a resolution of 2.29 Å. NO66 crystallized in space groupP31orP32, with unit-cell parametersa= 89.35,b = 89.35,c= 304.86 Å, α = β = 90, γ = 120°, and the crystal is likely to contain four molecules in the asymmetric unit.

scholarly journals Cloning, purification and preliminary crystallographic analysis of theHelicobacter pyloripseudaminic acid biosynthesisN-acetyltransferase PseH

2014 ◽  
Vol 70 (9) ◽  
pp. 1276-1279 ◽  
Author(s):  
Yu C. Liu ◽  
Abu I. Ud-Din ◽  
Anna Roujeinikova
Keyword(s):  
Crystallographic Analysis ◽  
Diffusion Method ◽  
X Ray Diffraction ◽  
Data Set ◽  
Cell Parameters ◽  
Nonulosonic Acid ◽  
Pseudaminic Acid ◽  
The Common ◽  
H Pylori ◽  
Development Of Gastric Cancer

Helicobacter pyloriinfection is the common cause of gastritis and duodenal and stomach ulcers, which have been linked to a higher risk of the development of gastric cancer. The motility that facilitates persistent infection requires functional flagella that are heavily glycosylated with 5,7-diacetamido-3,5,7,9-tetradeoxy-L-glycero-L-manno-nonulosonic acid (pseudaminic acid). Pseudaminic acid biosynthesis protein H (PseH) catalyzes the third step in its biosynthetic pathway, producing UDP-2,4-diacetamido-2,4,6-trideoxy-β-L-altropyranose. Crystals ofH. pyloriPseH have been grown by the hanging-drop vapour-diffusion method using diammonium tartrate as a precipitating agent. The crystals belonged to space groupI222 orI212121, with unit-cell parametersa= 107.8,b= 145.4,c= 166.3 Å. A complete X-ray diffraction data set has been collected to 2.5 Å resolution using cryocooling conditions and synchrotron radiation.

scholarly journals Cloning, expression, crystallization and crystallographic analysis of CouR fromRhodopseudomonas palustris

2015 ◽  
Vol 71 (11) ◽  
pp. 1416-1420 ◽  
Author(s):  
Chen Pan ◽  
Yong-lin Hu ◽  
Xiang-ning Jiang ◽  
Ying Gai
Keyword(s):  
Rhodopseudomonas Palustris ◽  
Crystallographic Analysis ◽  
Unit Cell ◽  
X Ray Diffraction ◽  
Unit Cell Parameters ◽  
Ligand Complex ◽  
Space Group ◽  
Cell Parameters ◽  
Dna Fragment

CouR fromRhodopseudomonas palustrisis a member of the MarR transcriptional regulator family. It regulates the expression of CouA and CouB, enzymes that are involved in the degradation ofp-coumarate.In vivo, CouR binds to a DNA fragment containing thecouABpromoter and suppresses the expression of CouA and CouB, while binding ofp-coumaroyl-CoA attenuates its affinity towards DNA and activates the expression of CouA and CouB. Here, the crystallization and X-ray diffraction analyses of CouR alone and in complex withp-coumaroyl-CoA are reported. Apo and ligand-complexed CouR crystals diffracted to 2.5 and 3.3 Å resolution, respectively. The crystals of apo CouR belonged to space groupP22121, with unit-cell parametersa= 62.78,b = 76.15,c = 87.38 Å, whereas the crystals of the CouR–ligand complex belonged to space groupP212121, with unit-cell parametersa= 61.37,b= 69.82,c = 70.32 Å. The crystals were predicted to contain two CouR molecules or CouR–ligand complexes per asymmetric unit.

scholarly journals Cloning, expression, purification, crystallization and X-ray crystallographic analysis of (S)-3-hydroxybutyryl-CoA dehydrogenase fromClostridium butyricum

2014 ◽  
Vol 70 (4) ◽  
pp. 485-488
Author(s):  
Eun-Jung Kim ◽  
Kyung-Jin Kim
Keyword(s):  
Crystallographic Analysis ◽  
Diffusion Method ◽  
Molecular Replacement ◽  
X Ray Diffraction ◽  
X Ray ◽  
Cell Parameters ◽  
Maximum Resolution ◽  
Replacement Method ◽  
Molecular Replacement Method ◽  
Crystal Volume

(S)-3-Hydroxybutyryl-CoA dehydrogenase fromClostridium butyricum(CbHBD) is an enzyme that catalyzes the second step in the biosynthesis ofn-butanol from acetyl-CoA by the reduction of acetoacetyl-CoA to 3-hydroxybutyryl-CoA. TheCbHBD protein was crystallized using the hanging-drop vapour-diffusion method in the presence of 2 Mammonium sulfate, 0.1 MCAPS pH 10.5, 0.2 Mlithium sulfate at 295 K. X-ray diffraction data were collected to a maximum resolution of 2.3 Å on a synchrotron beamline. The crystal belonged to space groupR3, with unit-cell parametersa=b= 148.5,c= 201.6 Å. With four molecules per asymmetric unit, the crystal volume per unit protein weight (VM) is 3.52 Å3 Da−1, which corresponds to a solvent content of approximately 65.04%. The structure was solved by the molecular-replacement method and refinement of the structure is in progress.

scholarly journals Purification, crystallization and preliminary X-ray crystallographic studies of Drep2 CIDE domain

2014 ◽  
Vol 70 (10) ◽  
pp. 1414-1417 ◽  
Author(s):  
Seung Mi Lee ◽  
Hyun Ho Park
Keyword(s):  
Fruit Fly ◽  
Diffusion Method ◽  
Amino Acid Residues ◽  
X Ray Diffraction ◽  
Hanging Drop ◽  
Unit Cell Parameters ◽  
Orthorhombic Space Group ◽  
X Ray ◽  
Cell Parameters ◽  
Protein Protein Interaction

Drep2 is a novel nuclease from the fruit fly that might have a similar function in apoptosis to DFF40 and DFF45, which are primary players in apoptotic DNA fragmentation. Drep2 contains a conserved CIDE domain of ∼90 amino-acid residues that is involved in protein–protein interaction. In this study, the Drep2 CIDE domain was purified and crystallized by the hanging-drop vapour-diffusion method. X-ray diffraction data were then collected to a resolution of 2.3 Å. The crystals were found to belong to the orthorhombic space groupP212121, with unit-cell parametersa= 50.28,b= 88.70,c= 113.37 Å.

scholarly journals Overexpression, purification, crystallization and preliminary X-ray crystallographic analysis of SF173 fromShigella flexneri

2015 ◽  
Vol 71 (1) ◽  
pp. 54-56 ◽  
Author(s):  
Ha-Neul Kim ◽  
Jeong-Gi An ◽  
Yoo-Sup Lee ◽  
Seung-Hyeon Seok ◽  
Hee-Seop Yoo ◽  
...  
Keyword(s):  
Anaerobic Bacterium ◽  
Shigella Flexneri ◽  
Hypothetical Protein ◽  
Crystallographic Analysis ◽  
Diffusion Method ◽  
X Ray Diffraction ◽  
Unit Cell Parameters ◽  
Gram Negative ◽  
X Ray ◽  
Cell Parameters

Shigella flexneriis a Gram-negative, anaerobic bacterium in the genusShigellathat can cause diarrhoea in humans. SF173, a hypothetical protein fromS. flexneri5a strain M90T, has been cloned, overexpressed, purified and crystallized as a part of laboratory-scale structural genomics project. The SF173 protein was crystallized using the sitting-drop vapour-diffusion method in the presence of 0.8 Msuccinic acid pH 7.0 at 293 K. Preliminary X-ray diffraction analysis revealed that the crystal diffracted to 1.47 Å resolution and belonged to space groupI432, with unit-cell parametersa=b=c= 110.245 Å.

scholarly journals Purification, crystallization and preliminary X-ray crystallographic analysis of glycosyltransferase-1 fromBacillus cereus

2014 ◽  
Vol 70 (9) ◽  
pp. 1228-1231 ◽  
Author(s):  
Yin-Cheng Hsieh ◽  
Hsi-Ho Chiu ◽  
Yen-Chieh Huang ◽  
Hoong-Kun Fun ◽  
Chia-Yu Lu ◽  
...  
Keyword(s):  
Small Molecules ◽  
Crystallographic Analysis ◽  
Diffusion Method ◽  
Uridine Diphosphate ◽  
X Ray Diffraction ◽  
X Ray ◽  
Cell Parameters ◽  
Biological Compounds ◽  
The Family ◽  
Diphosphate Glucose

Glycosyltransferases (GTs), which are distributed widely in various organisms, including bacteria, fungi, plants and animals, play a role in synthesizing biological compounds. Glycosyltransferase-1 fromBacillus cereus(BcGT-1), which is capable of transferring glucose to small molecules such as kaempferol and quercetin, has been identified as a member of the family 1 glycosyltransferases which utilize uridine diphosphate glucose (UDP-glucose) as the sugar donor.BcGT-1 (molecular mass 45.5 kDa) has been overexpressed, purified and crystallized using the hanging-drop vapour-diffusion method. According to X-ray diffraction ofBcGT-1 crystals to 2.10 Å resolution, the crystal belonged to space groupP1, with unit-cell parametersa= 54.56,b= 84.81,c= 100.12 Å, α = 78.36, β = 84.66, γ = 84.84°. Preliminary analysis indicates the presence of fourBcGT-1 molecules in the asymmetric unit with a solvent content of 50.27%.

scholarly journals Periplasmic form of dipeptidyl aminopeptidase IV fromPseudoxanthomonas mexicanaWO24: purification, kinetic characterization, crystallization and X-ray crystallographic analysis

2017 ◽  
Vol 73 (11) ◽  
pp. 601-606 ◽  
Author(s):  
Saori Roppongi ◽  
Chika Tateoka ◽  
Mayu Fujimoto ◽  
Ippei Iizuka ◽  
Saori Morisawa ◽  
...  
Keyword(s):  
Crystal Form ◽  
Crystallographic Analysis ◽  
Diffusion Method ◽  
X Ray Diffraction ◽  
X Ray ◽  
Cell Parameters ◽  
Replacement Method ◽  
Dpp Iv ◽  
Molecular Replacement Method ◽  
Dipeptidyl Aminopeptidase

Dipeptidyl aminopeptidase IV (DAP IV or DPP IV) fromPseudoxanthomonas mexicanaWO24 (PmDAP IV) preferentially cleaves substrate peptides with Pro or Ala at the P1 position [NH2-P2-P1(Pro/Ala)-P1′-P2′…]. For crystallographic studies, the periplasmic form of PmDAP IV was overproduced inEscherichia coli, purified and crystallized in complex with the tripeptide Lys-Pro-Tyr using the hanging-drop vapour-diffusion method. Kinetic parameters of the purified enzyme against a synthetic substrate were also determined. X-ray diffraction data to 1.90 Å resolution were collected from a triclinic crystal form belonging to space groupP1, with unit-cell parametersa= 88.66,b= 104.49,c = 112.84 Å, α = 67.42, β = 68.83, γ = 65.46°. Initial phases were determined by the molecular-replacement method usingStenotrophomonas maltophiliaDPP IV (PDB entry 2ecf) as a template and refinement of the structure is in progress.

scholarly journals Purification, crystallization and preliminary crystallographic analysis of the 16S rRNA methyltransferase RsmI fromEscherichia coli

2014 ◽  
Vol 70 (9) ◽  
pp. 1256-1259
Author(s):  
Mohan Zhao ◽  
Li Wang ◽  
Heng Zhang ◽  
Yuhui Dong ◽  
Yong Gong ◽  
...  
Keyword(s):  
16S Rrna ◽  
Crystallographic Analysis ◽  
Fine Tuning ◽  
Diffusion Method ◽  
X Ray Diffraction ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters ◽  
And Function ◽  
Decoding Fidelity

RsmI and RsmH are AdoMet-dependent methyltransferases that are responsible for the 2′-O-methylation andN4-methylation of C1402 ofEscherichia coli16S rRNA, respectively. Modification of this site has been found to play a role in fine-tuning the shape and function of the P-site to increase the decoding fidelity. It is interesting to study the mechanism by which C1402 can be methylated by both RsmI and RsmH. The crystal structure of RsmH in complex with AdoMet and cytidine has recently been determined and provided some implications forN4-methylation of this site. Here, the purification and crystallization of RsmI as well as its preliminary crystallographic analysis are reported. Co-crystallization of RsmI with AdoMet was carried out by the sitting-drop vapour-diffusion method and X-ray diffraction data were collected to 2.60 Å resolution on beamline 1W2B at BSRF. The crystal contained three molecules per asymmetric unit and belonged to space groupC2, with unit-cell parametersa= 121.9,b= 152.5,c= 54.2 Å, β = 93.4°.

scholarly journals Cloning, expression, purification, crystallization and X-ray crystallographic analysis of the (S)-3-hydroxybutyryl-CoA dehydrogenase PaaH1 fromRalstonia eutrophaH16

2014 ◽  
Vol 70 (7) ◽  
pp. 955-958 ◽  
Author(s):  
Jieun Kim ◽  
Kyung-Jin Kim
Keyword(s):  
Ralstonia Eutropha ◽  
Crystallographic Analysis ◽  
Diffusion Method ◽  
X Ray Diffraction ◽  
Hanging Drop ◽  
Dispersion Method ◽  
X Ray ◽  
Cell Parameters ◽  
Maximum Resolution ◽  
Crystal Volume

The (S)-3-hydroxybutyryl-CoA dehydrogenase PaaH1 fromRalstonia eutropha(RePaaH1) is an enzyme used in the biosynthesis ofn-butanol from acetyl-CoA by the reduction of acetoacetyl-CoA to (S)-3-hydroxybutyryl-CoA. TheRePaaH1 protein was crystallized using the hanging-drop vapour-diffusion method in the presence of 1.4 Mammonium sulfate, 0.1 Msodium cacodylate pH 6.0, 0.2 Msodium chloride at 295 K. X-ray diffraction data were collected to a maximum resolution of 2.6 Å on a synchrotron beamline. The crystal belonged to space groupP3221, with unit-cell parametersa=b= 135.4,c= 97.2 Å. With three molecules per asymmetric unit, the crystal volume per unit protein weight (VM) is 2.68 Å3 Da−1, which corresponds to a solvent content of approximately 54.1%. The structure was solved by the single-wavelength anomalous dispersion method and refinement of the structure is in progress.

scholarly journals The periplasmic sensing domain ofVibrio fischerichemoreceptor protein A (VfcA): cloning, purification and crystallographic analysis

2016 ◽  
Vol 72 (5) ◽  
pp. 382-385 ◽  
Author(s):  
Abu Iftiaf Md Salah Ud-Din ◽  
Anna Roujeinikova
Keyword(s):  
Vibrio Fischeri ◽  
Protein A ◽  
Crystallographic Analysis ◽  
Diffusion Method ◽  
Euprymna Scolopes ◽  
X Ray Diffraction ◽  
Hanging Drop ◽  
Data Set ◽  
X Ray ◽  
Cell Parameters

Flagella-mediated motility and chemotaxis towards nutrients are important characteristics ofVibrio fischerithat play a crucial role in the development of its symbiotic relationship with its Hawaiian squid hostEuprymna scolopes. TheV. fischerichemoreceptor A (VfcA) mediates chemotaxis toward amino acids. The periplasmic sensory domain of VfcA has been crystallized by the hanging-drop vapour-diffusion method using polyethylene glycol 3350 as a precipitating agent. The crystals belonged to space groupP1, with unit-cell parametersa = 39.9,b= 57.0,c= 117.0 Å, α = 88.9, β = 80.5, γ = 89.7°. A complete X-ray diffraction data set has been collected to 1.8 Å resolution using cryocooling conditions and synchrotron radiation.

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