Expression, crystallization and preliminary X-ray analysis of ligand-free human glutathione S-transferase M2–2

1998 ◽  
Vol 54 (3) ◽  
pp. 458-460 ◽  
Author(s):  
Larysa N. Patskovska ◽  
Alexander A. Fedorov ◽  
Yury V. Patskovsky ◽  
Steven C. Almo ◽  
Irving Listowsky
Keyword(s):  
Escherichia Coli ◽  
Active Site ◽  
Asymmetric Unit ◽  
Glutathione S Transferase ◽  
Crystal Forms ◽  
Space Group ◽  
X Ray ◽  
Ligand Free ◽  
Catalytically Active ◽  
Insight Into

Human glutathione-S-transferase M2–2 (hGSTM2–2) was expressed in Escherichia coli and purified by GSH-affinity chromatography. The recombinant enzyme and the protein isolated from human tissue were indistinguishable based on physicochemical, enzymatic and immunological criteria. The catalytically active dimeric hGSTM2–2 was crystallized without GSH or other active-site ligands in two crystal forms. Diffraction from form A crystals extends to 2.5 Å and is consistent with the space group P21 (a = 53.9, b = 81.5, c = 55.6 Å, β = 109.26 Å) with two monomers in the asymmetric unit. Diffraction from form B crystals extends to 3 Å and is consistent with a space group P212121 (a = 57.2, b = 80.7, c = 225.9 Å) with two dimers in the asymmetric unit. This is the first report of ligand-free mu-class GST crystals, and a comparison with liganded complexes will provide insight into the structural consequences of substrate binding which are thought to be important for catalysis.

Expression, purification, crystallization and preliminary X-ray data of Escherichia coli galactoside acetyltransferase

1999 ◽  
Vol 55 (11) ◽  
pp. 1955-1957 ◽  
Author(s):  
Xing-Guo Wang ◽  
Steven L. Roderick
Keyword(s):  
Escherichia Coli ◽  
Ammonium Sulfate ◽  
Asymmetric Unit ◽  
Acetyl Coa ◽  
Space Group ◽  
X Ray

Crystals of galactoside acetyltransferase from Escherichia coli have been prepared from solutions of ammonium sulfate containing acetyl-CoA. These crystals diffract to at least 2.7 Å resolution, belong to space group C2221 and contain one copy of the trimeric enzyme in the asymmetric unit.

scholarly journals Cloning, expression, purification and preliminary X-ray analysis of EstN2, a novel archaeal α/β-hydrolase fromCandidatusNitrososphaera gargensis

2014 ◽  
Vol 70 (10) ◽  
pp. 1394-1397 ◽  
Author(s):  
Heidi Kaljunen ◽  
Jennifer Chow ◽  
Wolfgang R. Streit ◽  
Jochen Mueller-Dieckmann
Keyword(s):  
Escherichia Coli ◽  
Lipase Activity ◽  
Lipolytic Activity ◽  
Recombinant Enzyme ◽  
X Rays ◽  
Asymmetric Unit ◽  
Heterologous Host ◽  
Space Group ◽  
X Ray

EstN2 is a novel α/β-hydrolase originating from the ammonia-oxidizing thaumarchaeonCandidatusNitrososphaera gargensis. The genome of the organism was sequenced and genes conferring putative lipolytic activity were amplified and cloned intoEscherichia colias a heterologous host. Through function-based screening, esterase and lipase activity was detected. A recombinant enzyme designated EstN2 was successfully expressed, purified and crystallized. The crystals belonged to space groupI2, with one molecule per asymmetric unit, and diffracted X-rays to 1.5 Å resolution.

scholarly journals Crystallization of importin α, the nuclear-import receptor

1999 ◽  
Vol 55 (2) ◽  
pp. 561-563 ◽  
Author(s):  
Trazel Teh ◽  
Tony Tiganis ◽  
Bostjan Kobe
Keyword(s):  
Nuclear Import ◽  
Sodium Citrate ◽  
Asymmetric Unit ◽  
Orthorhombic Space Group ◽  
Crystal Forms ◽  
Space Group ◽  
X Ray ◽  
Importin Α ◽  
Ph Conditions ◽  
Import Receptor

Crystals of recombinant importin α, the nuclear-import receptor, have been obtained at two different pH conditions by vapour diffusion using sodium citrate as precipitant and dithiothreitol as an additive. At pH 4–5, the crystals have the symmetry of the trigonal space group P3121 or P3221 (a = b = 78.0, c = 255.8 Å, γ = 120°); at pH 6–7, the crystals have the symmetry of the orthorhombic space group P212121 (a = 78.5, b = 89.7, c = 100.5 Å). In both cases, there is probably one molecule of importin α in the asymmetric unit. At least one of the crystal forms diffracts to a resolution higher than 3 Å using the laboratory X-ray source; the crystals are suitable for crystal structure determination.

scholarly journals Crystallization and preliminary X-ray analysis of a ribokinase fromVibrio choleraeO395

2014 ◽  
Vol 70 (8) ◽  
pp. 1098-1102 ◽  
Author(s):  
Rakhi Paul ◽  
Madhumita Dandopath Patra ◽  
Ramanuj Banerjee ◽  
Udayaditya Sen
Keyword(s):  
Escherichia Coli ◽  
Polyethylene Glycol ◽  
Catalytic Mechanism ◽  
Empty Space ◽  
Adenosine Diphosphate ◽  
Asymmetric Unit ◽  
Solvent Content ◽  
X Ray ◽  
Polyethylene Glycol 6000 ◽  
Insight Into

Ribokinase (RK) is one of the principal enzymes in carbohydrate metabolism, catalyzing the reaction of D-ribose and adenosine triphosphate to produce ribose-5-phosphate and adenosine diphosphate (ADP). To provide further insight into the catalytic mechanism, therbsKgene fromVibrio choleraeO395 encoding ribokinase was cloned and the protein was overexpressed inEscherichia coliBL21 (DE3) and purified using Ni2+–NTA affinity chromatography. Crystals ofV. choleraeRK (Vc-RK) and of its complex with ribose and ADP were grown in the presence of polyethylene glycol 6000 and diffracted to 3.4 and 1.75 Å resolution, respectively. Analysis of the diffraction data showed that both crystals possess symmetry consistent with space groupP1. In the Vc-RK crystals, 16 molecules in the asymmetric unit were arranged in a spiral fashion, leaving a large empty space inside the crystal, which is consistent with its high Matthews coefficient (3.9 Å3 Da−1) and solvent content (68%). In the Vc-RK co-crystals four molecules were located in the asymmetric unit with a Matthews coefficient of 2.4 Å3 Da−1, corresponding to a solvent content of 50%.

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.

Crystallization and preliminary X-ray study of two crystal forms of Klebsiella oxytoca diol dehydratase–cyanocobalamin complex

1999 ◽  
Vol 55 (4) ◽  
pp. 907-909 ◽  
Author(s):  
Jun Masuda ◽  
Tetsuya Yamaguchi ◽  
Takamasa Tobimatsu ◽  
Tetsuo Toraya ◽  
Kyoko Suto ◽  
...  
Keyword(s):  
Klebsiella Oxytoca ◽  
Crystal Form ◽  
Unit Cell ◽  
Unit Cell Parameters ◽  
Crystal Forms ◽  
Space Group ◽  
X Ray ◽  
Cell Parameters ◽  
Diol Dehydratase ◽  
Replacement Method

Two crystal forms of Klebsiella oxytoca diol dehydratase complexed with cyanocobalamin have been obtained and preliminary crystallographic experiments have been performed. The crystals belong to two different space groups, depending on the crystallization conditions. One crystal (form I) belongs to space group P212121 with unit-cell parameters a = 76.2, b = 122.3, c = 209.6 Å, and diffracts to 2.2 Å resolution using an X-ray beam from a synchrotron radiation source. The other crystal (form II) belongs to space group P21 with unit-cell parameters a = 75.4, b = 132.7, c = 298.8 Å, β = 91.9°, and diffracts to 3.0 Å resolution. For the purpose of structure determination, a heavy-atom derivative search was carried out and some mercuric derivatives were found to be promising. Structure analysis by the multiple isomorphous replacement method is now under way.

Sur la structure cristalline des dimethyl-1,1 germa-1 (et sila-1) cycloundécanediols-6,7

1975 ◽  
Vol 53 (23) ◽  
pp. 3596-3598 ◽  
Author(s):  
François Brisse ◽  
Aviva Battat ◽  
Jean-Claude Richer ◽  
Pierre Mazerolles ◽  
Alfreda Faucher
Keyword(s):  
Asymmetric Unit ◽  
Space Group ◽  
X Ray ◽  
Independent Molecules

1,1-Dimethyl-1-germa (and-1-sila) -6,7-cycloundecanediol (C12H26O2Ge and C12H26O2Si) are isostructural as established by their X-ray powder patterns. The dimensions of the triclinic cells are as follows: for the silicon derivative, a = 10.53, b = 12.45, and c = 12.43 Å, α = 81.5°, β = 67.0°, and γ = 76.3°; for the germanium derivative, a = 10.56, b = 12.50, and c = 12.58 Å, α = 82.1°, β = 67.8°, and γ = 76.1°. If the space group is [Formula: see text] there will be two independent molecules in each asymmetric unit.

scholarly journals The three-dimensional structure of a class-Pi glutathione S-transferase complexed with glutathione: the active-site hydration provides insights into the reaction mechanism

1998 ◽  
Vol 333 (3) ◽  
pp. 811-816 ◽  
Author(s):  
Antonio PÁRRAGA ◽  
Isabel GARCÍA-SÁEZ ◽  
Sinead B. WALSH ◽  
Timothy J. MANTLE ◽  
Miquel COLL
Keyword(s):  
Conformational Changes ◽  
Active Site ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Water Molecules ◽  
X Ray Diffraction ◽  
Glutathione S Transferase ◽  
X Ray ◽  
The Right

The structure of mouse liver glutathione S-transferase P1-1 complexed with its substrate glutathione (GSH) has been determined by X-ray diffraction analysis. No conformational changes in the glutathione moiety or in the protein, other than small adjustments of some side chains, are observed when compared with glutathione adduct complexes. Our structure confirms that the role of Tyr-7 is to stabilize the thiolate by hydrogen bonding and to position it in the right orientation. A comparison of the enzyme–GSH structure reported here with previously described structures reveals rearrangements in a well-defined network of water molecules in the active site. One of these water molecules (W0), identified in the unliganded enzyme (carboxymethylated at Cys-47), is displaced by the binding of GSH, and a further water molecule (W4) is displaced following the binding of the electrophilic substrate and the formation of the glutathione conjugate. The possibility that one of these water molecules participates in the proton abstraction from the glutathione thiol is discussed.

scholarly journals Diosgenone: a secondP21polymorph

2012 ◽  
Vol 68 (8) ◽  
pp. o2358-o2358 ◽  
Author(s):  
María-Guadalupe Hernández Linares ◽  
Gabriel Guerrero-Luna ◽  
Sylvain Bernès ◽  
Marcos Flores-Alamo ◽  
María A. Fernández-Herrera
Keyword(s):  
Relative Orientation ◽  
Asymmetric Unit ◽  
Space Group ◽  
X Ray ◽  
Therapeutic Alternative ◽  
Structure Report ◽  
Independent Molecules

Diosgenone [(20S,22R,25R)-spirost-4-en-3-one, C27H40O3] has been proposed as a new therapeutic alternative for the treatment of malaria. The first X-ray structure report for diosgenone was by Piroet al.[(2002).Z. Naturforsch. Teil C,57, 947–950] in the space groupP21(Z′ = 2). We now report a new polymorph in the same space group, with two molecules in the asymmetric unit. Both molecules have similar conformations, characterized by a skewed envelopeAring, which contains the C=C bond conjugated with the ketone functionality at C3. The dimorphism results from a modification of the relative orientation of the molecules in the asymmetric unit: two independent molecules were arranged antiparallel in the Piro report, while they are parallel in the present determination.

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