Crystallization and preliminary crystallographic study of triple-helical DNA

2000 ◽  
Vol 56 (1) ◽  
pp. 104-105 ◽  
Author(s):  
Zong-Jin Han ◽  
Sangkee Rhee ◽  
Keliang Liu ◽  
H. Todd Miles ◽  
David R. Davies
Keyword(s):  
Single Crystals ◽  
Unit Cell ◽  
The Other ◽  
Diffusion Method ◽  
Triplex Dna ◽  
Asymmetric Unit ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters ◽  
Crystallographic Study

Single crystals of d(CTCCTSCCGCGCG)·d(CGCGCGGAG) have been grown by the vapor-diffusion method using 2-methyl-2,4-pentanediol as a precipitant. The crystals are tetragonal, space group P42, with unit-cell parameters a = b = 53.8, c = 43.1 Å, and diffract to 1.8 Å resolution at a synchrotron X-ray beamline. In the crystal, the asymmetric unit contains one copy of the construct. The two halves of the structure are related by non-crystallographic twofold symmetry. These observations are consistent with the conclusion that the sequences of the 12-mer and 9-mer oligonucleotides form a duplex DNA at one end and a triplex DNA at the other end.

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 diffraction analysis of hydroxynitrile lyase from cassava (Manihot esculenta)

1999 ◽  
Vol 55 (4) ◽  
pp. 904-906 ◽  
Author(s):  
Hanspeter Lauble ◽  
Klaas Decanniere ◽  
Harald Wajant ◽  
Siegfried Förster ◽  
Franz Effenberger
Keyword(s):  
Unit Cell ◽  
Diffusion Method ◽  
Asymmetric Unit ◽  
Unit Cell Parameters ◽  
Orthorhombic Space Group ◽  
Data Set ◽  
Hydroxynitrile Lyase ◽  
Space Group ◽  
X Ray ◽  
Cell Parameters

Hydroxynitrile lyase from M. esculenta (cassava) was crystallized in two different crystal forms by the hanging-drop vapour-diffusion method. Crystals of form I were obtained from a mixture of polyethylene glycol 8000 and 2-methyl-2,4-pentanediol, and belong to the tetragonal space group P41212 or its enantiomorph P43212, with unit-cell parameters a  =  b  =  105.9, c  =  188.9 Å and with two molecules in the asymmetric unit. These crystals diffract to 2.9 Å with conventional X-ray sources and beyond 2.1 Å resolution with synchrotron radiation. The crystals are relatively sensitive to radiation damage and conditions for flash-cooling the crystals have been established. A complete native data set has been collected up to 2.2 Å resolution. Crystal form II has been obtained at pH 5.6 using lithium sulfate as a precipitant. The crystals apparently belong to the orthorhombic space group P21212, with unit-cell parameters a = 117.52, b = 127.09 and c = 78.08 Å, have two molecules in the asymmetric unit and diffract to beyond 2.0 Å resolution. A complete native data set has been collected to 2.2 Å resolution.

Analysis of an industrial production suspension ofBacillus lentussubtilisin crystals by powder diffraction: a powerful quality-control tool

2014 ◽  
Vol 70 (4) ◽  
pp. 1115-1123 ◽  
Author(s):  
Christian G. Frankaer ◽  
Olga V. Moroz ◽  
Johan P. Turkenburg ◽  
Stein I. Aspmo ◽  
Majbritt Thymark ◽  
...  
Keyword(s):  
Quality Control ◽  
Single Crystals ◽  
Powder Diffraction ◽  
Unit Cell ◽  
Unit Cell Parameters ◽  
Space Group ◽  
X Ray ◽  
Cell Parameters ◽  
Quality Control Tool ◽  
Control Tool

A microcrystalline suspension ofBacillus lentussubtilisin (Savinase) produced during industrial large-scale production was analysed by X-ray powder diffraction (XRPD) and X-ray single-crystal diffraction (MX). XRPD established that the bulk microcrystal sample representative of the entire production suspension corresponded to space groupP212121, with unit-cell parametersa= 47.65,b= 62.43,c= 75.74 Å, equivalent to those for a known orthorhombic crystal form (PDB entry 1ndq). MX using synchrotron beamlines at the Diamond Light Source with beam dimensions of 20 × 20 µm was subsequently used to study the largest crystals present in the suspension, with diffraction data being collected from two single crystals (∼20 × 20 × 60 µm) to resolutions of 1.40 and 1.57 Å, respectively. Both structures also belonged to space groupP212121, but were quite distinct from the dominant form identified by XRPD, with unit-cell parametersa= 53.04,b = 57.55,c= 71.37 Å anda= 52.72,b= 57.13,c= 65.86 Å, respectively, and refined toR= 10.8% andRfree= 15.5% and toR= 14.1% andRfree= 18.0%, respectively. They are also different from any of the forms previously reported in the PDB. A controlled crystallization experiment with a highly purified Savinase sample allowed the growth of single crystals of the form identified by XRPD; their structure was solved and refined to a resolution of 1.17 Å with anRof 9.2% and anRfreeof 11.8%. Thus, there are at least three polymorphs present in the production suspension, albeit with the 1ndq-like microcrystals predominating. It is shown how the two techniques can provide invaluable and complementary information for such a production suspension and it is proposed that XRPD provides an excellent quality-control tool for such suspensions.

scholarly journals Cloning, purification and preliminary X-ray crystallographic analysis of the OmpA-like domain of peptidoglycan-associated lipoprotein fromAcinetobacter baumannii

2012 ◽  
Vol 68 (11) ◽  
pp. 1351-1353 ◽  
Author(s):  
Jung Hyun Song ◽  
Woo Cheol Lee ◽  
Jeong Soon Park ◽  
Seung Il Kim ◽  
Je Chul Lee ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Acinetobacter Baumannii ◽  
Crystallographic Analysis ◽  
Diffusion Method ◽  
Asymmetric Unit ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters ◽  
Vapour Diffusion ◽  
Strain Bl21

Peptidoglycan-associated lipoprotein (Pal) is one component of the Tol–Pal system that is involved in maintaining the integrity and stability of the outer membrane. The C-terminal OmpA-like domain of Pal interacts noncovalently with peptidoglycan. In this study, the OmpA-like domain of Pal fromAcinetobacter baumanniiwas overexpressed inEscherichia colistrain BL21 (DE3), purified and crystallized using the vapour-diffusion method. A native crystal diffracted to 1.4 Å resolution and belonged to space groupP61orP65, with unit-cell parametersa=b= 72.58,c= 44.65 Å, a calculated Matthews coefficient of 2.64 Å3 Da−1and one molecule per asymmetric unit.

scholarly journals Purification, crystallization and preliminary X-ray crystallographic studies of Rv3705c fromMycobacterium tuberculosis

2014 ◽  
Vol 70 (8) ◽  
pp. 1090-1092
Author(s):  
Feifei Lu ◽  
Feng Gao ◽  
Honglin Li ◽  
Weimin Gong ◽  
Lin Zhou ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Unit Cell ◽  
Diffusion Method ◽  
Unit Cell Parameters ◽  
Space Group ◽  
X Ray ◽  
Cell Parameters ◽  
Vapour Diffusion

The conserved protein Rv3705c fromMycobacterium tuberculosishas been cloned, expressed, purified and crystallized by the sitting-drop vapour-diffusion method using PEG 3350 as a precipitant. The Rv3705c crystals exhibited space groupP6122 orP6522, with unit-cell parametersa=b= 198.0,c= 364.1 Å, α = β = 90, γ = 120°, and diffracted to a resolution of 3.3 Å.

scholarly journals Crystallization and preliminary X-ray analysis of the NAD+-reducing [NiFe] hydrogenase fromHydrogenophilus thermoluteolusTH-1

2015 ◽  
Vol 71 (1) ◽  
pp. 96-99 ◽  
Author(s):  
Midori Taketa ◽  
Hanae Nakagawa ◽  
Mao Habukawa ◽  
Hisao Osuka ◽  
Kiyohito Kihira ◽  
...  
Keyword(s):  
Diffusion Method ◽  
Asymmetric Unit ◽  
Dispersion Method ◽  
Unit Cell Parameters ◽  
Solvent Content ◽  
X Ray ◽  
Cell Parameters ◽  
Vapour Diffusion ◽  
Anomalous Signal ◽  
Th 1

NAD+-reducing [NiFe] hydrogenases catalyze the oxidoreduction of dihydrogen concomitant with the interconversion of NAD+and NADH. Here, the isolation, purification and crystallization of the NAD+-reducing [NiFe] hydrogenase fromHydrogenophilus thermoluteolusTH-1 are reported. Crystals of the NAD+-reducing [NiFe] hydrogenase were obtained within one week from a solution containing polyethylene glycol using the sitting-drop vapour-diffusion method and micro-seeding. The crystal diffracted to 2.58 Å resolution and belonged to space groupC2, with unit-cell parametersa= 131.43,b= 189.71,c= 124.59 Å, β = 109.42°. Assuming the presence of two NAD+-reducing [NiFe] hydrogenase molecules in the asymmetric unit,VMwas calculated to be 2.2 Å3 Da−1, which corresponds to a solvent content of 43%. Initial phases were determined by the single-wavelength anomalous dispersion method using the anomalous signal from the Fe atoms.

scholarly journals Crystallization and preliminary X-ray analysis of Rv1674c fromMycobacterium tuberculosis

2015 ◽  
Vol 71 (3) ◽  
pp. 354-357 ◽  
Author(s):  
Jincheng Li ◽  
Xudong Wang ◽  
Weimin Gong ◽  
Chunyan Niu ◽  
Min Zhang
Keyword(s):  
Diffusion Method ◽  
Dna Binding Domain ◽  
Asymmetric Unit ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters ◽  
Protein Molecules ◽  
Vapour Diffusion ◽  
Transcription Regulators ◽  
Interdomain Interaction

Adaptations to hypoxia play an important role inMycobacterium tuberculosispathogenesis. Rv0324, which contains an HTH DNA-binding domain and a rhodanese domain, is one of the key transcription regulators in response to hypoxia.M. tuberculosisRv1674c is a homologue of Rv0324. To understand the interdomain interaction and regulation of the HTH domain and the rhodanese domain, recombinant Rv1674c protein was purified and crystallized by the vapour-diffusion method. The crystals diffracted to 2.25 Å resolution. Preliminary diffraction analysis suggests that the crystals belonged to space groupP3121 orP3221, with unit-cell parametersa=b= 67.8,c= 174.5 Å, α = β = 90, γ = 120°. The Matthews coefficient was calculated to be 2.44 Å3 Da−1, assuming that the crystallographic asymmetric unit contains two protein molecules.

scholarly journals Neutron crystallographic study of heterotrimeric glutamine amidotransferase CAB

2019 ◽  
Vol 75 (3) ◽  
pp. 193-196
Author(s):  
Long Li ◽  
Motoyasu Adachi ◽  
Jian Yu ◽  
Koji Kato ◽  
Akira Shinoda ◽  
...  
Keyword(s):  
Neutron Diffraction ◽  
Diffraction Data ◽  
Active Sites ◽  
Neutron Diffraction Data ◽  
Asymmetric Unit ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters ◽  
Crystallographic Study ◽  
Glutamine Amidotransferase

Heterotrimeric glutamine amidotransferase CAB (GatCAB) possesses an ammonia-self-sufficient mechanism in which ammonia is produced and used in the inner complex by GatA and GatB, respectively. The X-ray structure of GatCAB revealed that the two identified active sites of GatA and GatB are markedly distant, but are connected in the complex by a channel of 30 Å in length. In order to clarify whether ammonia is transferred through this channel in GatCAB by visualizing ammonia, neutron diffraction studies are indispensable. Here, GatCAB crystals were grown to approximate dimensions of 2.8 × 0.8 × 0.8 mm (a volume of 1.8 mm3) with the aid of a polymer using microseeding and macroseeding processes. Monochromatic neutron diffraction data were collected using the neutron single-crystal diffractometer BIODIFF at the Heinz Maier-Leibnitz Zentrum, Germany. The GatCAB crystals belonged to space group P212121, with unit-cell parameters a = 74.6, b = 94.5, c = 182.5 Å and with one GatCAB complex (molecular mass 119 kDa) in the asymmetric unit. This study represented a challenge in current neutron diffraction technology.

scholarly journals Crystallization and preliminary X-ray diffraction analysis ofBacillus subtilisYwfE, anL-amino-acid ligase

2012 ◽  
Vol 68 (2) ◽  
pp. 203-206 ◽  
Author(s):  
Takeo Tsuda ◽  
Tomomi Suzuki ◽  
Shuichi Kojima
Keyword(s):  
Amino Acid ◽  
Diffusion Method ◽  
Dependent Manner ◽  
Asymmetric Unit ◽  
X Ray Diffraction ◽  
Hanging Drop ◽  
Unit Cell Parameters ◽  
Data Set ◽  
X Ray ◽  
Cell Parameters

Bacillus subtilisYwfE, an L-amino-acid ligase, catalyzes the formation of an α-dipeptide from L-amino acids in an ATP-dependent manner. In order to elucidate the substrate-recognition mode and the reaction mechanism of this ligase, native and selenomethionine-derivatized (SeMet) crystals of YwfE in the presence of ADP, MgCl2and the dipeptide L-Ala-L-Gln were obtained using the hanging-drop vapour-diffusion method. These crystals diffracted to 1.9 and 2.8 Å resolution, respectively. Preliminary SAD phase calculations using the data set from the SeMet crystal suggested that the crystal belonged to the hexagonal space groupP6522, with unit-cell parametersa=b= 90.85,c = 250.31 Å, and contained one molecule in the asymmetric unit with a solvent content of 57.3%.

Expression, purification, crystallization and preliminary X-ray analysis of the κ-carrageenase from Pseudoalteromonas carrageenovora

1999 ◽  
Vol 55 (4) ◽  
pp. 918-920 ◽  
Author(s):  
Gurvan Michel ◽  
Tristan Barbeyron ◽  
Didier Flament ◽  
Thierry Vernet ◽  
Bernard Kloareg ◽  
...  
Keyword(s):  
Polyethylene Glycol ◽  
Unit Cell ◽  
Recombinant Form ◽  
Diffusion Method ◽  
Unit Cell Parameters ◽  
Space Group ◽  
X Ray ◽  
Cell Parameters ◽  
Vapour Diffusion

A recombinant form of His-tagged κ-carrageenase from Pseudoalteromonas carrageenovora has been expressed, purified and crystallized. Crystals have been obtained by the vapour-diffusion method using polyethylene glycol (Mr = 4000) as a precipitant. These crystals belong to the space group P212121, with unit-cell parameters a  =  58.2, b  =  62.8, c  =  77.9 Å, and diffract to 2.2 Å resolution on a rotating-anode X-ray source.

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