scholarly journals Crystallographic studies of SarV, a global regulator fromStaphylococcus aureus

2015 ◽  
Vol 71 (8) ◽  
pp. 1038-1041
Author(s):  
Yang Song ◽  
Fan Zhang ◽  
Xu Li ◽  
Jianye Zang ◽  
Xuan Zhang
Keyword(s):  
Heavy Atom ◽  
Size Exclusion ◽  
Monoclinic Space Group ◽  
Molecular Replacement ◽  
Global Regulator ◽  
X Ray Diffraction ◽  
X Ray ◽  
Cell Parameters ◽  
Exclusion Chromatography ◽  
Heavy Atom Method

SarV, a member of the SarA protein family, is a global transcriptional regulator which has been reported to be involved in the regulation of autolysis inStaphylococcus aureus. In this study, SarV fromS. aureuswas successfully cloned, expressed, purified and crystallized. X-ray diffraction data were collected to 2.10 Å resolution. The crystals of SarV belonged to the monoclinic space groupP21, with unit-cell parametersa= 36.40,b= 119.64,c= 66.80 Å, α = γ = 90, β = 98.75°. The Matthews coefficient and the solvent content were estimated to be 2.57 Å3 Da−1and 52%, respectively, suggesting the presence of four molecules in the asymmetric unit. The results of size-exclusion chromatography (SEC) indicated thatS. aureusSarV exists as a homodimer in solution. Unfortunately, the structure cannot be solved by molecular replacement because of the low sequence identity ofS. aureusSarV to known structures. Further phase determination by selenomethionine single-wavelength anomalous dispersion (SAD) and the heavy-atom method is in progress.

scholarly journals Purification, crystallization and preliminary X-ray diffraction studies of UDP-glucose:tetrahydrobiopterin α-glucosyltransferase (BGluT) fromSynechococcussp. PCC 7942

2014 ◽  
Vol 70 (2) ◽  
pp. 203-205
Author(s):  
Asaithambi Killivalavan ◽  
Ningning Zhuang ◽  
Young Shik Park ◽  
Kon Ho Lee
Keyword(s):  
Size Exclusion ◽  
Diffusion Method ◽  
Monoclinic Space Group ◽  
X Ray Diffraction ◽  
Dispersion Method ◽  
X Ray ◽  
Cell Parameters ◽  
Exclusion Chromatography ◽  
Multi Wavelength ◽  
Pcc 7942

A UDP-glucose:tetrahydrobiopterin α-glucosyltransferase (BGluT) enzyme was discovered in the cyanobacteriumSynechococcussp. PCC 7942 which transfers a glucose moiety from UDP-glucose to tetrahydrobiopterin (BH4). BGluT protein was overexpressed with selenomethionine labelling for structure determination by the multi-wavelength anomalous dispersion method. The BGluT protein was purified by nickel-affinity and size-exclusion chromatography. It was then crystallized by the hanging-drop vapour-diffusion method using a well solution consisting of 0.1 Mbis-tris pH 5.5, 19%(w/v) polyethylene glycol 3350 with 4%(w/v) D(+)-galactose as an additive. X-ray diffraction data were collected to 1.99 Å resolution using a synchrotron-radiation source. The crystals belonged to the monoclinic space groupC2, with unit-cell parametersa= 171.35,b= 77.99,c = 53.77 Å, β = 90.27°.

scholarly journals Purification, crystallization and preliminary X-ray diffraction analysis of ArsH fromSynechocystissp. strain PCC 6803

2014 ◽  
Vol 70 (4) ◽  
pp. 497-500 ◽  
Author(s):  
Xiao Zhang ◽  
Xi-Mei Xue ◽  
Yu Yan ◽  
Jun Ye
Keyword(s):  
Sinorhizobium Meliloti ◽  
Size Exclusion ◽  
Diffusion Method ◽  
Flavin Mononucleotide ◽  
Molecular Replacement ◽  
X Ray Diffraction ◽  
X Ray ◽  
Cell Parameters ◽  
Exclusion Chromatography ◽  
Pcc 6803

ArsH is an NADPH-dependent flavin mononucleotide reductase and is frequently encoded as part of anarsoperon. The function of thearsHgene remains to be characterized. Crystallization and structural studies may contribute to elucidating the specific biological function of ArsH associated with arsenic resistance. ArsH fromSynechocystissp. strain PCC 6803 was overproduced, purified and crystallized. Crystals were obtained by the sitting-drop vapour-diffusion method. Diffraction data were collected and processed to a resolution of 1.6 Å. The crystals belonged to the tetragonal space groupI4122, with unit-cell parametersa=b= 127.94,c= 65.86 Å and one molecule in the asymmetric unit. Size-exclusion chromatography and molecular-replacement results showed that the ArsH formed a tetramer. Further structural analysis and comparison with ArsH fromSinorhizobium melilotiwill provide information about the oligomerization of ArsH.

scholarly journals Cloning, expression, purification, crystallization and preliminary X-ray diffraction of a lysine-specific permease fromPseudomonas aeruginosa

2014 ◽  
Vol 70 (10) ◽  
pp. 1362-1367 ◽  
Author(s):  
Emmanuel Nji ◽  
Dianfan Li ◽  
Declan A. Doyle ◽  
Martin Caffrey
Keyword(s):  
Metal Ion ◽  
Cell Volume Regulation ◽  
Ion Concentration ◽  
Size Exclusion ◽  
Substrate Selectivity ◽  
X Ray Diffraction ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters ◽  
Exclusion Chromatography

The prokaryotic lysine-specific permease (LysP) belongs to the amino acid–polyamine–organocation (APC) transporter superfamily. In the cell, members of this family are responsible for the uptake and recycling of nutrients, for the maintenance of a constant internal ion concentration and for cell volume regulation. The detailed mechanism of substrate selectivity and transport of L-lysine by LysP is not understood. A high-resolution crystal structure would enormously facilitate such an understanding. To this end, LysP fromPseudomonas aeruginosawas recombinantly expressed inEscherichia coliand purified to near homogeneity by immobilized metal ion-affinity chromatography (IMAC) and size-exclusion chromatography (SEC). Hexagonal- and rod-shaped crystals were obtained in the presence of L-lysine and the L-lysine analogue L-4-thialysine by vapour diffusion and diffracted to 7.5 Å resolution. The diffraction data were indexed in space groupP21, with unit-cell parametersa= 169.53,b= 169.53,c= 290.13 Å, γ = 120°.

scholarly journals A putative siderophore-interacting protein from the marine bacteriumShewanella frigidimarinaNCIMB 400: cloning, expression, purification, crystallization and X-ray diffraction analysis

2016 ◽  
Vol 72 (9) ◽  
pp. 667-671 ◽  
Author(s):  
Inês B. Trindade ◽  
Bruno M. Fonseca ◽  
Pedro M. Matias ◽  
Ricardo O. Louro ◽  
Elin Moe
Keyword(s):  
Iron Acquisition ◽  
Crystallographic Analysis ◽  
Monoclinic Space Group ◽  
Molecular Replacement ◽  
X Ray Diffraction ◽  
Interacting Protein ◽  
X Ray ◽  
Cell Parameters ◽  
Unit Structure ◽  
Shewanella Frigidimarina

Siderophore-binding proteins (SIPs) perform a key role in iron acquisition in multiple organisms. In the genome of the marine bacteriumShewanella frigidimarinaNCIMB 400, the gene tagged as SFRI_RS12295 encodes a protein from this family. Here, the cloning, expression, purification and crystallization of this protein are reported, together with its preliminary X-ray crystallographic analysis to 1.35 Å resolution. The SIP crystals belonged to the monoclinic space groupP21, with unit-cell parametersa= 48.04,b= 78.31,c= 67.71 Å, α = 90, β = 99.94, γ = 90°, and are predicted to contain two molecules per asymmetric unit. Structure determination by molecular replacement and the use of previously determined ∼2 Å resolution SIP structures with ∼30% sequence identity as templates are ongoing.

Crystal and Molecular Structure of Dithiocyanato(triphenylarsine)mercury(II)

1975 ◽  
Vol 53 (22) ◽  
pp. 3383-3387 ◽  
Author(s):  
Joseph Hubert ◽  
André L. Beauchamp ◽  
Roland Rivest
Keyword(s):  
Molecular Structure ◽  
Least Squares ◽  
Coordination Number ◽  
Diffraction Data ◽  
Crystal And Molecular Structure ◽  
Heavy Atom ◽  
Monoclinic Space Group ◽  
X Ray Diffraction ◽  
X Ray ◽  
Heavy Atom Method

The crystal and molecular structure of dithiocyanato(triphenylarsine)mercury(II) has been determined from X-ray diffraction data. The crystals are monoclinic, space group P21/c, with a = 10.290(7), b = 21.199(23), c = 10.719(7) Å, β = 112.00(2)°, and Z = 4. The structure has been solved by the heavy-atom method and refined by block-diagonal least-squares calculations. The agreement factor R obtained for 2607 'observed' reflections is 0.030. The crystal consists of single molecules. The 'characteristic' coordination number of mercury is three, with two sulfur and one arsenic atoms at the apexes of a triangle. The nitrogen atoms of the thiocyanate groups are at 2.67 and 2.74 Å from the adjoining mercury atoms and therefore link the different molecules together.

Crystallization and Preliminary X-ray Diffraction Analysis of Cold-Active β-Galactosidase from Arthrobacter sp. C2-2

2005 ◽  
Vol 70 (1) ◽  
pp. 124-132 ◽  
Author(s):  
Hana Petroková ◽  
Eva Vondráčková ◽  
Tereza Skálová ◽  
Jan Dohnálek ◽  
Petra Lipovová ◽  
...  
Keyword(s):  
Structure Refinement ◽  
Monoclinic Space Group ◽  
Molecular Replacement ◽  
Phase Problem ◽  
X Ray Diffraction ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters ◽  
Psychrotrophic Bacteria ◽  
Cold Active

β-Galactosidase from psychrotrophic bacteria strainArthrobactersp. C2-2 catalyzes cleavage of β-D-galactosyl moieties from β-D-galactosides and is interesting for its activity at low temperatures. Various types of crystals with dimensions of up to 0.8 mm were obtained and X-ray diffraction data up to 1.9 Å were collected. The crystals belong to the monoclinic space groupP21with unit-cell parametersa= 140.1 Å,b= 205.7 Å,c= 140.5 Å and β = 102.3°. The enzyme (molecular weight of a monomer is 111 kDa) forms hexamers in the crystal structure (one hexamer per asymmetric unit). The phase problem was solved by molecular replacement. Structure refinement is in progress.

scholarly journals Purification, crystallization and preliminary X-ray diffraction analysis of theEscherichia colicommon pilus chaperone EcpB

2015 ◽  
Vol 71 (6) ◽  
pp. 676-679 ◽  
Author(s):  
James A. Garnett ◽  
Mamou Diallo ◽  
Steve J. Matthews
Keyword(s):  
Heavy Atom ◽  
Three Dimensional ◽  
Solid Surfaces ◽  
Dimensional Structure ◽  
Molecular Replacement ◽  
X Ray Diffraction ◽  
X Ray ◽  
Cell Parameters ◽  
The Common ◽  
First Time

Pili are key cell-surface components that allow the attachment of bacteria to both biological and abiotic solid surfaces, whilst also mediating interactions between themselves. InEscherichia coli, the common pilus (Ecp) belongs to an alternative chaperone–usher (CU) pathway that plays a major role in both early biofilm formation and host-cell adhesion. The chaperone EcpB is involved in the biogenesis of the filament, which is composed of EcpA and EcpD. Initial attempts at crystallizing EcpB using natively purified protein from the bacterial periplasm were not successful; however, after the isolation of EcpB under denaturing conditions and subsequent refolding, crystals were obtained at pH 8.0 using the sitting-drop method of vapour diffusion. Diffraction data have been processed to 2.4 Å resolution. These crystals belonged to the trigonal space groupP3121 orP3221, with unit-cell parametersa=b= 62.65,c= 121.14 Å and one monomer in the asymmetric unit. Molecular replacement was unsuccessful, but selenomethionine-substituted protein and heavy-atom derivatives are being prepared for phasing. The three-dimensional structure of EcpB will provide invaluable information on the subtle mechanistic differences in biogenesis between the alternative and classical CU pathways. Furthermore, this is the first time that this refolding strategy has been used to purify CU chaperones, and it could be implemented in similar systems where it has not been possible to obtain highly ordered crystals.

The crystal and molecular structure of di-μ-phenylthiolatobis[trichloro(dimethyl sulfide) tungsten(IV) (W—W)]

1983 ◽  
Vol 61 (12) ◽  
pp. 2809-2812 ◽  
Author(s):  
P. Michael Boorman ◽  
Joanne M. Ball ◽  
Kelly J. Moynihan ◽  
Vikram D. Patel ◽  
John F. Richardson
Keyword(s):  
Molecular Structure ◽  
Bond Length ◽  
Dimethyl Sulfide ◽  
Crystal And Molecular Structure ◽  
Heavy Atom ◽  
Monoclinic Space Group ◽  
X Ray Diffraction ◽  
X Ray ◽  
Sulfide Ligands ◽  
Heavy Atom Method

The complex (Me2S)Cl3W(μ-SPh)2WCl3(SMe2), 1, has been isolated as one product of the 1:1 reaction between WCl4(Me2S)2 and SiMe3(SPh) in CH2Cl2 solution. A single crystal X-ray diffraction study shows that the molecule has the relatively unusual edge-shared bioctahedral structure, with a W—W bond length of 2.759(1) Å. The dimethyl sulfide ligands occupy positions trans to one another in the equatorial mean plane of the molecule, which has two-fold symmetry imposed on it. The structure was solved by the heavy atom method and refined to R = 0.044 and Rw = 0.058 for 2001 reflections. Crystals of 1 are monoclinic, space group C2/c, with a = 17.445(4), b = 12.594(2), c = 11.509(3) Å, β = 91.22(1)°, and Z = 4.

Crystallization and preliminary X-ray diffraction studies of a Bowman–Birk inhibitor from Vigna unguiculata seeds

1999 ◽  
Vol 55 (11) ◽  
pp. 1920-1922 ◽  
Author(s):  
K. N. Rao ◽  
S. S. Hegde ◽  
R. J. Lewis ◽  
C. G. Suresh
Keyword(s):  
Vigna Unguiculata ◽  
Diffusion Method ◽  
Monoclinic Space Group ◽  
Molecular Replacement ◽  
X Ray Diffraction ◽  
X Ray ◽  
Cell Parameters ◽  
Replacement Method ◽  
Molecular Replacement Method ◽  
Citrate Phosphate

A Bowman–Birk type trypsin/chymotrypsin inhibitor isolated from Vigna unguiculata seeds has been crystallized. Crystals were grown using the vapour-diffusion method at pH 4.0 using citrate/phosphate as a buffer and 30% saturated ammonium sulfate as precipitant. The crystals belonged to the monoclinic space group P21, with unit-cell parameters a = 32.4, b = 61.8, c = 32.9 Å, β = 114.5°. The Matthews coefficient calculated assuming two molecules in the asymmetric unit was 1.95 Å3 Da−1, which corresponds to a 37% solvent content. X-ray data were collected to 2.5 Å resolution from a flash-frozen crystal. The structure was solved using the molecular-replacement method using tracy soybean inhibitor structure (PDB entry 1pi2) as a model.

scholarly journals Purification and crystallographic analysis of a FAD-dependent halogenase fromStreptomycessp. JCM9888

2015 ◽  
Vol 71 (8) ◽  
pp. 972-976 ◽  
Author(s):  
Yanqun Zhao ◽  
Baohua Yan ◽  
Ting Yang ◽  
Jian Jiang ◽  
Heng Wei ◽  
...  
Keyword(s):  
Model Building ◽  
Structure Refinement ◽  
Crystallographic Analysis ◽  
Monoclinic Space Group ◽  
Molecular Replacement ◽  
X Ray Diffraction ◽  
X Ray ◽  
Cell Parameters ◽  
Hexahistidine Tag ◽  
Crystallization Experiments

A new FAD (flavin adenine dinucleotide)-dependent halogenase HalY fromStreptomycessp. JCM9888 was reported to be involved in the regioselective halogenation of adenine. HalY is a variant B FAD-dependent halogenase that is most similar to the halogenase PltA involved in pyoluteorin biosynthesis. This study reports the overexpression and purification of HalY with an N-terminal hexahistidine tag, followed by crystallization experiments and X-ray crystallographic analysis. HalY was purified as a monomer in solution and crystallized to give X-ray diffraction to a resolution of 1.7 Å. The crystal belonged to the monoclinic space groupP21, with unit-cell parametersa= 41.4,b= 113.4,c= 47.6 Å, α = γ = 90, β = 107.4°, and contained one monomer of HalY in the asymmetric unit, with a calculated Matthews coefficient of 2.3 Å3 Da−1and a solvent content of 46%. The structure of the halogenase CndH was used as a search model in molecular replacement to obtain the initial model of HalY. Manual model building and structure refinement of HalY are in progress.

Sign in / Sign up

Export Citation Format

Share Document