scholarly journals Expression, purification, crystallization and preliminary X-ray crystallographic analysis of the histone-like HU protein fromSpiroplasma melliferumKC3

2015 ◽  
Vol 71 (1) ◽  
pp. 24-27 ◽  
Author(s):  
Konstantin Boyko ◽  
Marina Gorbacheva ◽  
Tatiana Rakitina ◽  
Dmitry Korzhenevskiy ◽  
Anna Vanyushkina ◽  
...  
Keyword(s):  
Crystallographic Analysis ◽  
Molecular Replacement ◽  
Dna Compaction ◽  
Hu Protein ◽  
Replacement Method ◽  
Hu Proteins ◽  
Diffusion Technique ◽  
Associated Proteins ◽  
Molecular Replacement Method ◽  
Spiroplasma Melliferum

HU proteins belong to the nucleoid-associated proteins (NAPs) that are involved in vital processes such as DNA compaction and reparation, gene transcriptionetc.No data are available on the structures of HU proteins from mycoplasmas. To this end, the HU protein from the parasitic mycoplasmaSpiroplasma melliferumKC3 was cloned, overexpressed inEscherichia coliand purified to homogeneity. Prismatic crystals of the protein were obtained by the vapour-diffusion technique at 4°C. The crystals diffracted to 1.36 Å resolution (the best resolution ever obtained for a HU protein). The diffraction data were indexed in space groupC2 and the structure of the protein was solved by the molecular-replacement method with one monomer per asymmetric unit.

scholarly journals Crystallization and preliminary X-ray crystallographic analysis of PBPD2 fromListeria monocytogenes

2014 ◽  
Vol 70 (4) ◽  
pp. 535-537 ◽  
Author(s):  
Hyung Jin Cha ◽  
Jae-Hee Jeong ◽  
Yeon-Gil Kim
Keyword(s):  
Biosynthetic Pathway ◽  
Crystallographic Analysis ◽  
Diffusion Method ◽  
Low Molecular Weight ◽  
Molecular Replacement ◽  
Orthorhombic Space Group ◽  
Penicillin Binding Proteins ◽  
Cell Parameters ◽  
Replacement Method ◽  
Molecular Replacement Method

Penicillin-binding proteins (PBPs), which mediate the peptidoglycan biosynthetic pathway in the bacterial cell wall, have been intensively investigated as a target for the design of antibiotics. In this study, PBPD2, a low-molecular-weight PBP encoded bylmo2812fromListeria monocytogenes, was overexpressed inEscherichia coli, purified and crystallized at 295 K using the sitting-drop vapour-diffusion method. The crystal belonged to the primitive orthorhombic space groupP212121, with unit-cell parametersa= 37.7,b= 74.7,c= 75.1 Å, and diffracted to 1.55 Å resolution. There was one molecule in the asymmetric unit. The preliminary structure was determined by the molecular-replacement method.

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.

Crystallization and preliminary structural studies of Scilla campanulata lectin complexed with α1–6 mannobiose

1998 ◽  
Vol 54 (1) ◽  
pp. 90-92 ◽  
Author(s):  
Lisa M Wright ◽  
Stephen D. Wood ◽  
Colin D. Reynolds ◽  
Pierre J. Rizkallah ◽  
Anthony K. Allen
Keyword(s):  
Molecular Replacement ◽  
X Rays ◽  
Synchrotron Source ◽  
Replacement Method ◽  
Cell Dimensions ◽  
Molecular Replacement Method ◽  
Hanging Drop Method ◽  
Unit Cell Dimensions ◽  
Antiretroviral Activity ◽  
Image Plate System

Recent work has shown that Scilla campanulata agglutinin from bluebell bulbs has a strong affinity for α(1,3)- and α(1,6)-linked mannosyl residues and possesses moderate antiretroviral activity. This lectin has been crystallized by the hanging-drop method of vapour diffusion complexed with the disaccharide mannose-α1,6-D-mannose. The crystals are in the space group P21212 with unit-cell dimensions a = 70.63, b = 92.79 and c = 47.25 Å, and with a dimer in the asymmetric unit. The crystals diffract X-rays to beyond 1.5 Å resolution at 277 K and are stable in an X-ray beam. Data to 1.6 Å resolution have been collected using a MAR image-plate system at a synchrotron source and the structure of the complex has been solved by the molecular replacement method.

scholarly journals Cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of AerF fromMicrocystis aeruginosa, a putative reductase participating in aeruginosin biosynthesis

2015 ◽  
Vol 71 (4) ◽  
pp. 466-470 ◽  
Author(s):  
Ruyi Ding ◽  
Cui Xu ◽  
Xu Chen ◽  
Mengyun Bao ◽  
Xiaoting Qiu
Keyword(s):  
Diffraction Analysis ◽  
Biosynthetic Pathway ◽  
Molecular Replacement ◽  
X Ray Diffraction ◽  
Antithrombotic Activity ◽  
X Ray ◽  
Cell Parameters ◽  
Maximum Resolution ◽  
Replacement Method ◽  
Molecular Replacement Method

The 2-carboxy-6-hydroxyoctahydroindole moiety is an essential residue for the antithrombotic activity of aeruginosins, which are a class of cyanobacteria-derived bioactive linear tetrapeptides. The biosynthetic pathway of the 2-carboxy-6-hydroxyoctahydroindole moiety has not yet been resolved. AerF was indicated to be involved in the biosynthesis of the 2-carboxy-6-hydroxyoctahydroindole moiety. This study reports the cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of AerF fromMicrocystis aeruginosawith a C-terminal His6tag. The crystal diffracted to a maximum resolution of 1.38 Å and belonged to the tetragonal space groupP4322, with unit-cell parametersa=b= 101.581,c= 116.094 Å. The calculated Matthews coefficient and solvent content of the crystal were 2.47 Å3 Da−1and 50.32%, respectively. The initial model of the structure was obtained by the molecular-replacement method and refinement of the structure is in progress.

Clarithromycin form I determined by synchrotron X-ray powder diffraction

2012 ◽  
Vol 68 (2) ◽  
pp. o41-o44 ◽  
Author(s):  
Shuji Noguchi ◽  
Keiko Miura ◽  
Sadahiro Fujiki ◽  
Yasunori Iwao ◽  
Shigeru Itai
Keyword(s):  
Powder Diffraction ◽  
Crystal Packing ◽  
Diffraction Method ◽  
Molecular Replacement ◽  
The Novel ◽  
Intermolecular Hydrogen Bonds ◽  
Rietveld Refinements ◽  
Replacement Method ◽  
Metastable Form ◽  
Molecular Replacement Method

The structure of the metastable form I polymorph of the macrolide antibiotic clarithromycin, C38H69NO13, was determined by a powder diffraction method using synchrotron radiation. The space group of form I isP21212. The initial model was determined by a molecular replacement method using the structure of clarithromycin form 0 as a search model, and the final structure was obtained through Rietveld refinements. In the form I crystal structure, the clarithromycin molecules are aligned parallel along theaaxis in a head-to-tail manner with intermolecular hydrogen bonds between the hydroxy O atoms. The dimethylamine groups of the clarithromycin molecule interdigitate between neighbouring head-to-tail clarithromycin alignments. The novel crystal packing found in form I provides a mechanism that describes the transformation of form 0 to form I.

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