scholarly journals Crystallographic analysis of RsmA, a ribosomal RNA small subunit methyltransferase A fromStaphylococcus aureus

2015 ◽  
Vol 71 (8) ◽  
pp. 1063-1066 ◽  
Author(s):  
Yang Liu ◽  
Yuwei Zhu ◽  
Maikun Teng ◽  
Xu Li
Keyword(s):  
Staphylococcus Aureus ◽  
Ribosomal Rna ◽  
Small Subunit ◽  
Crystallographic Analysis ◽  
Asymmetric Unit ◽  
X Ray Diffraction ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters ◽  
Rotation Function

RsmA, a ribosomal RNA small subunit methyltransferase fromStaphylococcus aureus, catalyzes theN6methylation of adenine in 16S rRNA. In this study, RsmA fromStaphylococcus aureuswas cloned, expressed, purified and crystallized. The crystal belonged to space groupC2, with unit-cell parametersa= 84.38,b= 157.76,c= 96.50 Å, β = 95.04°. X-ray diffraction data were collected to a resolution of 3.2 Å. The self-rotation function and the Matthews coefficient suggested the presence of two molecules in the asymmetric unit.

scholarly journals Overproduction, crystallization and preliminary X-ray crystallographic analysis ofEscherichia colitRNAN6-threonylcarbamoyladenosine dehydratase

2014 ◽  
Vol 70 (11) ◽  
pp. 1517-1520 ◽  
Author(s):  
Sunmin Kim ◽  
Keon Young Kim ◽  
Jeong Kuk Park ◽  
Byung Il Lee ◽  
Yun-Gon Kim ◽  
...  
Keyword(s):  
Crystallographic Analysis ◽  
X Rays ◽  
Asymmetric Unit ◽  
X Ray Diffraction ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters ◽  
E Coli ◽  
Protein Mass ◽  
Crystal Volume

Escherichia colitRNAN6-threonylcarbamoyladenosine dehydratase (TcdA), previously called CsdL or YgdL, was overproduced and purified fromE. coliand crystallized using polyethylene glycol 3350 as a crystallizing agent. X-ray diffraction data were collected to 2.70 Å resolution under cryoconditions using synchrotron X-rays. The crystals belonged to space groupP21, with unit-cell parametersa= 65.4,b= 96.8,c= 83.3 Å, β = 111.7°. According to the Matthews coefficient, the asymmetric unit may contain up to four subunits of the monomeric protein, with a crystal volume per protein mass (VM) of 2.12 Å3 Da−1and 42.1% solvent content.

scholarly journals Expression, purification and preliminary crystallographic analysis ofMycobacterium tuberculosisCysQ, a phosphatase involved in sulfur metabolism

2014 ◽  
Vol 70 (6) ◽  
pp. 750-753 ◽  
Author(s):  
Anna I. Erickson ◽  
Reta D. Sarsam ◽  
Andrew J. Fisher
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Sulfur Metabolism ◽  
Crystallographic Analysis ◽  
Transfer Reactions ◽  
Asymmetric Unit ◽  
X Ray Diffraction ◽  
Unit Cell Parameters ◽  
Orthorhombic Space Group ◽  
X Ray ◽  
Cell Parameters

CysQ is part of the sulfur-activation pathway that dephosphorylates 3′-phosphoadenosine 5′-monophosphate (PAP) to regenerate adenosine 5′-monophosphate (AMP) and free phosphate. PAP is the product of sulfate-transfer reactions from sulfotransferases that use the universal sulfate donor 3′-phosphoadenosine 5′-phosphosulfate (PAPS). In some organisms PAP is also the product of PAPS reductases that reduce sulfate from PAPS to sulfite. CysQ fromMycobacterium tuberculosis, which plays an important role in the biosynthesis of sulfated glycoconjugates, was successfully purified and crystallized in 24% PEG 1500, 20% glycerol. X-ray diffraction data were collected to 1.7 Å resolution using a synchrotron-radiation source. Crystals grew in the orthorhombic space groupP212121, with unit-cell parametersa= 40.3,b= 57.9,c= 101.7 Å and with one monomer per asymmetric unit.

scholarly journals Crystallization and preliminary X-ray crystallographic analysis of ribosome assembly factors: the Rpf2–Rrs1 complex

2014 ◽  
Vol 70 (12) ◽  
pp. 1649-1652 ◽  
Author(s):  
Nozomi Asano ◽  
Akiyoshi Nakamura ◽  
Keisuke Komoda ◽  
Koji Kato ◽  
Isao Tanaka ◽  
...  
Keyword(s):  
Ribosomal Proteins ◽  
Ribosome Biogenesis ◽  
Crystallographic Analysis ◽  
Asymmetric Unit ◽  
X Ray Diffraction ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters ◽  
Essential Proteins ◽  
Unit Structure

Rpf2 and Rrs1 are essential proteins for ribosome biogenesis. These proteins form a complex (the Rpf2-subcomplex) with 5S rRNA and two ribosomal proteins (L5 and L11). This complex is recruited to the ribosome precursor (the 90S pre-ribosome). This recruitment is necessary for the maturation of 25S rRNA. Genetic depletion of Rpf2 and Rrs1 results in accumulation of the 25S rRNA precursor. In this study, Rpf2 and Rrs1 fromAspergillus nidulanswere co-overexpressed inEscherichia coli, purified and crystallized. Subsequent analysis revealed that these crystals contained the central core region of the complex consisting of both N-terminal domains. X-ray diffraction data were collected to 2.35 Å resolution. Preliminary analysis revealed that the crystals belonged to space groupP212121, with unit-cell parametersa= 54.1,b= 123.3,c = 133.8 Å. There are two complexes in the asymmetric unit. Structure determination using selenomethionine-labelled protein is in progress.

scholarly journals Ectodomain of plasmodesmata-localized protein 5 inArabidopsis: expression, purification, crystallization and crystallographic analysis

2017 ◽  
Vol 73 (9) ◽  
pp. 532-535 ◽  
Author(s):  
Xiaocui Wang ◽  
Peiyan Zhu ◽  
Shanshan Qu ◽  
Jie Zhao ◽  
Prashant K. Singh ◽  
...  
Keyword(s):  
Expression System ◽  
Cell Movement ◽  
Crystallographic Analysis ◽  
Asymmetric Unit ◽  
X Ray Diffraction ◽  
Unit Cell Parameters ◽  
Solvent Content ◽  
X Ray ◽  
Cell Parameters ◽  
Crystal Content

Plasmodesmata-localized protein 5 (PDLP5) is a cysteine-rich receptor-like protein which is localized on the plasmodesmata ofArabidopsis thaliana. Overexpression of PDLP5 can reduce the permeability of the plasmodesmata and further affect the cell-to-cell movement of viruses and macromolecules in plants. The ectodomain of PDLP5 contains two DUF26 domains; however, the function of these domains is still unknown. Here, the ectodomain of PDLP5 fromArabidopsiswas cloned and overexpressed using an insect expression system and was then purified and crystallized. X-ray diffraction data were collected to 1.90 Å resolution and were indexed in space groupP1, with unit-cell parametersa= 41.9,b= 48.1,c = 62.2 Å, α = 97.3, β = 103.1, γ = 99.7°. Analysis of the crystal content indicated that there are two molecules in the asymmetric unit, with a Matthews coefficient of 2.51 Å3 Da−1and a solvent content of 50.97%.

scholarly journals Crystallization and X-ray diffraction analysis of native and selenomethionine-substituted PhyH-DI fromBacillussp. HJB17

2017 ◽  
Vol 73 (11) ◽  
pp. 607-611
Author(s):  
Fang Lu ◽  
Bei Zhang ◽  
Yong Liu ◽  
Ying Song ◽  
Gangxing Guo ◽  
...  
Keyword(s):  
Unit Cell ◽  
Diffusion Method ◽  
Data Sets ◽  
Asymmetric Unit ◽  
X Ray Diffraction ◽  
Unit Cell Parameters ◽  
Solvent Content ◽  
Space Group ◽  
X Ray ◽  
Cell Parameters

Phytases are phosphatases that hydrolyze phytates to less phosphorylatedmyo-inositol derivatives and inorganic phosphate. β-Propeller phytases, which are very diverse phytases with improved thermostability that are active at neutral and alkaline pH and have absolute substrate specificity, are ideal substitutes for other commercial phytases. PhyH-DI, a β-propeller phytase fromBacillussp. HJB17, was found to act synergistically with other single-domain phytases and can increase their efficiency in the hydrolysis of phytate. Crystals of native and selenomethionine-substituted PhyH-DI were obtained using the vapour-diffusion method in a condition consisting of 0.2 Msodium chloride, 0.1 MTris pH 8.5, 25%(w/v) PEG 3350 at 289 K. X-ray diffraction data were collected to 3.00 and 2.70 Å resolution, respectively, at 100 K. Native PhyH-DI crystals belonged to space groupC121, with unit-cell parametersa = 156.84,b = 45.54,c = 97.64 Å, α = 90.00, β = 125.86, γ = 90.00°. The asymmetric unit contained two molecules of PhyH-DI, with a corresponding Matthews coefficient of 2.17 Å3 Da−1and a solvent content of 43.26%. Crystals of selenomethionine-substituted PhyH-DI belonged to space groupC2221, with unit-cell parametersa = 94.71,b= 97.03,c= 69.16 Å, α = β = γ = 90.00°. The asymmetric unit contained one molecule of the protein, with a corresponding Matthews coefficient of 2.44 Å3 Da−1and a solvent content of 49.64%. Initial phases for PhyH-DI were obtained from SeMet SAD data sets. These data will be useful for further studies of the structure–function relationship of PhyH-DI.

scholarly journals Purification, crystallization and preliminary crystallographic analysis of a 4-thiouridine synthetase–RNA complex

2013 ◽  
Vol 69 (4) ◽  
pp. 421-424 ◽  
Author(s):  
Peter-Thomas Naumann ◽  
Charles T. Lauhon ◽  
Ralf Ficner
Keyword(s):  
Diffraction Data ◽  
Thermotoga Maritima ◽  
Crystallographic Analysis ◽  
X Ray Diffraction ◽  
Unit Cell Parameters ◽  
Orthorhombic Space Group ◽  
X Ray ◽  
Cell Parameters ◽  
Dependent Modification ◽  
Rna Complex

The sulfurtransferase 4-thiouridine synthetase (ThiI) is involved in the ATP-dependent modification of U8 in tRNA. ThiI fromThermotoga maritimawas cloned, overexpressed and purified. A complex comprising ThiI and a truncated tRNA was prepared and crystallized, and X-ray diffraction data were collected to a resolution of 3.5 Å. The crystals belonged to the orthorhombic space groupP212121, with unit-cell parametersa= 102.9,b= 112.8,c= 132.8 Å.

scholarly journals Crystallization and preliminary X-ray crystallographic analysis of human myotubularin-related protein 1

2015 ◽  
Vol 71 (3) ◽  
pp. 261-265 ◽  
Author(s):  
Seoung Min Bong ◽  
Seung Won Yang ◽  
Ji-Woong Choi ◽  
Seung Jun Kim ◽  
Byung Il Lee
Keyword(s):  
Escherichia Coli ◽  
Polyethylene Glycol ◽  
Synchrotron Radiation ◽  
Crystallographic Analysis ◽  
Asymmetric Unit ◽  
Related Protein ◽  
Unit Cell Parameters ◽  
Solvent Content ◽  
X Ray ◽  
Cell Parameters

Myotubularin-related protein 1 is a phosphatase that dephosphorylates phospholipids such as phosphatidylinositol 3-phosphate or phosphatidylinositol 3,5-bisphosphate. In this study, human MTMR1 was overexpressed inEscherichia coli, purified and crystallized at 277 K using polyethylene glycol 20 000 as a precipitant. Diffraction data were collected to 2.0 Å resolution using synchrotron radiation. The crystals belonged to space groupP1, with unit-cell parametersa= 67.219,b= 96.587,c= 97.581 Å, α = 87.597, β = 86.072, γ = 77.327°. Assuming the presence of four molecules in the asymmetric unit, the calculated Matthews coefficient value was 2.61 Å3 Da−1and the corresponding solvent content was 52.9%.

scholarly journals Expression, purification, crystallization and preliminary X-ray crystallographic analysis of Enpp6

2014 ◽  
Vol 70 (6) ◽  
pp. 794-799 ◽  
Author(s):  
Junko Morita ◽  
Kazuki Kato ◽  
Emiko Mihara ◽  
Ryuichiro Ishitani ◽  
Junichi Takagi ◽  
...  
Keyword(s):  
Catalytic Domain ◽  
Crystallographic Analysis ◽  
X Ray Diffraction ◽  
Unit Cell Parameters ◽  
Data Set ◽  
X Ray ◽  
Cell Parameters ◽  
Choline Metabolism ◽  
Protein Molecules ◽  
Membrane Bound

Enpp (ectonucleotide phosphodiesterase/pyrophosphatase) 6 is a membrane-bound glycoprotein that hydrolyzes choline-containing compounds such as lysophosphatidylcholine and glycerophosphorylcholine, and presumably participates in choline metabolism. The catalytic domain of mouse Enpp6 was expressed in HEK293T cells, purified using the TARGET tag/P20.1-Sepharose system and crystallized. An X-ray diffraction data set was collected to 1.8 Å resolution. The crystal belonged to space groupP1, with unit-cell parametersa= 63.7,b= 68.8,c= 69.7 Å, α = 60.6, β = 87.0, γ = 68.1°. Assuming the presence of two protein molecules per asymmetric unit, the solvent content was estimated to be 49.5%.

Preparation and preliminary X-ray diffraction studies of a new crystal form of L-asparaginase from Escherichia coli

1999 ◽  
Vol 55 (9) ◽  
pp. 1616-1617
Author(s):  
I. Polikarpov ◽  
R. T. de Oliveira ◽  
J. Abrahão-Neto
Keyword(s):  
Escherichia Coli ◽  
Synchrotron Radiation ◽  
Crystal Form ◽  
Asymmetric Unit ◽  
Chemotherapeutic Drug ◽  
X Ray Diffraction ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters ◽  
Synchrotron Radiation Diffraction

L-Asparaginase is an enzyme which hydrolyzes asparagine to produce aspartic acid and ammonia. It is an effective chemotherapeutic drug, especially in the treatment of acute lymphoblastic leukaemia in children. The enzyme from Escherichia coli was crystallized in a new crystal form with space group C2, unit-cell parameters a = 76.3 (0), b = 134.6 (2), c = 64.8 (7) Å, β = 110.5 (1)° and a dimer in the asymmetric unit. Synchrotron-radiation diffraction data have been collected to 1.95 Å resolution.

scholarly journals Purification, crystallization and X-ray crystallographic analysis of human RAB11(S20V), a constitutively active GTP-binding form

2015 ◽  
Vol 71 (10) ◽  
pp. 1247-1250 ◽  
Author(s):  
Chang Min Kim ◽  
Jae Young Choi ◽  
Jong Hwan Yoon ◽  
Hyun Ho Park
Keyword(s):  
Crystallographic Analysis ◽  
X Ray Diffraction ◽  
Unit Cell Parameters ◽  
Gtp Hydrolysis ◽  
X Ray ◽  
Cell Parameters ◽  
Gtp Binding ◽  
His Tag ◽  
Hydrolysis Activity ◽  
Constitutively Active

RAB11, a member of the Ras superfamily of small G proteins, is involved in the regulation of vesicle trafficking during endosome recycling. Substitution of Ser20 by Val20 in Rab11 [RAB11(S20V)] inhibits its GTP hydrolysis activity and produces a constitutively active GTP-binding form. In this study, the RAB11(S20V) mutant was overexpressed inEscherichia coliwith an engineered C-terminal His tag. RAB11(S20V) was then purified to homogeneity and was crystallized at 293 K. X-ray diffraction data were collected to a resolution of 2.4 Å from a crystal belonging to space groupI4, with unit-cell parametersa = 74.11,b= 74.11,c= 149.44 Å. The asymmetric unit was estimated to contain two molecules of RAB11(S20V).

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