Crystallization and preliminary diffraction studies of a truncated form of a novel protease from spores of Bacillus megaterium

2000 ◽  
Vol 56 (1) ◽  
pp. 70-72 ◽  
Author(s):  
Karthe Ponnuraj ◽  
Stephen Kelly ◽  
Claudio Nessi ◽  
Peter Setlow ◽  
Mark J. Jedrzejas
Keyword(s):  
Synchrotron Radiation ◽  
Bacillus Megaterium ◽  
Diffusion Method ◽  
Monoclinic Space Group ◽  
X Rays ◽  
Unit Cell Parameters ◽  
Truncated Form ◽  
Cell Parameters ◽  
Peg 4000 ◽  
Spore Proteins

During germination of spores of Bacillus species, a novel protease termed GPR initiates the degradation of a group of small acid-soluble spore proteins which protect the dormant spore's DNA from damage. Trypsin digestion of the zymogen of B. megaterium GPR removes ∼15 kDa from the C-terminal end of the 46 kDa zymogen subunit, leaving a 30 kDa subunit. Single crystals of this truncated form of GPR have been obtained by the vapor-diffusion method using PEG 4000 as a precipitating agent. The crystals belong to the monoclinic space group P21, with unit-cell parameters a = 67.99, b = 105.34, c = 108.63 Å, β = 95.68°. The cryofrozen crystals diffract X-rays to about 3.3 Å using synchrotron radiation.

Crystallization and preliminary X-ray diffraction studies of undecaprenyl diphosphate synthase from Micrococcus luteus B-P 26

1999 ◽  
Vol 55 (9) ◽  
pp. 1606-1607 ◽  
Author(s):  
Masahiro Fujihashi ◽  
Naoto Shimizu ◽  
Yuan-Wei Zhang ◽  
Tanetoshi Koyama ◽  
Kunio Miki
Keyword(s):  
Micrococcus Luteus ◽  
Crystal Structure Analysis ◽  
Diffusion Method ◽  
Monoclinic Space Group ◽  
Lithium Sulfate ◽  
X Rays ◽  
X Ray Diffraction ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters

Undecaprenyl diphosphate synthase from Micrococcus luteus B-P 26, one of the Z-prenyl chain-elongating enzymes, was crystallized using the sitting-drop vapour-diffusion method with ammonium sulfate and lithium sulfate as precipitants. The crystals belong to the monoclinic space group C2, with unit-cell parameters a = 127.2, b = 60.2, c = 75.7 Å, β = 105.6°. The crystals diffract X-rays to at least 2.2 Å resolution using synchrotron radiation and are suitable for high-resolution crystal structure analysis.

scholarly journals Crystallization and preliminary X-ray crystallographic analysis of the ArsM arsenic(III)S-adenosylmethionine methyltransferase

2010 ◽  
Vol 66 (9) ◽  
pp. 1050-1052 ◽  
Author(s):  
Kavitha Marapakala ◽  
A. Abdul Ajees ◽  
Jie Qin ◽  
Banumathi Sankaran ◽  
Barry P. Rosen
Keyword(s):  
Crystallographic Analysis ◽  
Hazardous Substances ◽  
Diffusion Method ◽  
Monoclinic Space Group ◽  
Hanging Drop ◽  
Unit Cell Parameters ◽  
Priority List ◽  
X Ray ◽  
Cell Parameters ◽  
Environmental Toxin

Arsenic is the most ubiquitous environmental toxin and carcinogen and consequently ranks first on the Environmental Protection Agency's Superfund Priority List of Hazardous Substances. It is introduced primarily from geochemical sources and is acted on biologically, creating an arsenic biogeocycle. A common biotransformation is methylation to monomethylated, dimethylated and trimethylated species. Methylation is catalyzed by the ArsM (or AS3MT) arsenic(III)S-adenosylmethionine methyltransferase, an enzyme (EC 2.1.1.137) that is found in members of every kingdom from bacteria to humans. ArsM from the thermophilic algaCyanidioschyzonsp. 5508 was expressed, purified and crystallized. Crystals were obtained by the hanging-drop vapor-diffusion method. The crystals belonged to the monoclinic space groupC2, with unit-cell parametersa= 84.85,b= 46.89,c= 100.35 Å, β = 114.25° and one molecule in the asymmetric unit. Diffraction data were collected at the Advanced Light Source and were processed to a resolution of 1.76 Å.

scholarly journals Expression, purification and crystallization of a plant polyketide cyclase fromCannabis sativa

2015 ◽  
Vol 71 (12) ◽  
pp. 1470-1474 ◽  
Author(s):  
Xinmei Yang ◽  
Takashi Matsui ◽  
Takahiro Mori ◽  
Futoshi Taura ◽  
Hiroshi Noguchi ◽  
...  
Keyword(s):  
Catalytic Mechanism ◽  
Cannabis Sativa ◽  
Crystallographic Analysis ◽  
Diffusion Method ◽  
X Rays ◽  
Unit Cell Parameters ◽  
Cell Parameters ◽  
Sequence Identity ◽  
Aldol Cyclization ◽  
Function Relationship

Plant polyketides are a structurally diverse family of natural products. In the biosynthesis of plant polyketides, the construction of the carbocyclic scaffold is a key step in diversifying the polyketide structure. Olivetolic acid cyclase (OAC) fromCannabis sativaL. is the only known plant polyketide cyclase that catalyzes the C2–C7 intramolecular aldol cyclization of linear pentyl tetra-β-ketide-CoA to generate olivetolic acid in the biosynthesis of cannabinoids. The enzyme is also thought to belong to the dimeric α+β barrel (DABB) protein family. However, because of a lack of functional analysis of other plant DABB proteins and low sequence identity with the functionally distinct bacterial DABB proteins, the catalytic mechanism of OAC has remained unclear. To clarify the intimate catalytic mechanism of OAC, the enzyme was overexpressed inEscherichia coliand crystallized using the vapour-diffusion method. The crystals diffracted X-rays to 1.40 Å resolution and belonged to space groupP3121 orP3221, with unit-cell parametersa=b= 47.3,c= 176.0 Å. Further crystallographic analysis will provide valuable insights into the structure–function relationship and catalytic mechanism of OAC.

scholarly journals Pteridine glycosyltransferase from Chlorobium tepidum: crystallization and X-ray analysis

2017 ◽  
Vol 73 (11) ◽  
pp. 629-634
Author(s):  
Asaithambi Killivalavan ◽  
Young Shik Park ◽  
Kon Ho Lee
Keyword(s):  
Diffusion Method ◽  
Chlorobium Tepidum ◽  
Monoclinic Space Group ◽  
Hanging Drop ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters ◽  
Chromatographic Techniques ◽  
Multiple Wavelength ◽  
Reservoir Solution

The pteridine glycosyltransferase (PGT) found in Chlorobium tepidum (CtPGT) catalyzes the conversion of L-threo-tetrahydrobiopterin to 1-O-(L-threo-biopterin-2′-yl)-β-N-acetylglucosamine using UDP-N-acetylglucosamine. The gene for CtPGT was cloned, and selenomethionine-derivatized protein was overexpressed and purified using various chromatographic techniques. The protein was crystallized by the hanging-drop vapour-diffusion method using 0.24 M triammonium citrate pH 7.0, 14%(w/v) PEG 3350 as a reservoir solution. Multiple-wavelength anomalous diffraction data were collected to 2.15 Å resolution from a single CtPGT crystal. The crystal belonged to the monoclinic space group C2, with unit-cell parameters a = 189.61, b = 79.98, c = 105.92 Å, β = 120.5°.

scholarly journals Structures of ruthenium-modified Pseudomonas aeruginosa azurin and [Ru(2,2′-bipyridine)2(imidazole)2]SO4·10H2O

1999 ◽  
Vol 55 (2) ◽  
pp. 379-385 ◽  
Author(s):  
Salem Faham ◽  
Michael W. Day ◽  
William B. Connick ◽  
Brian R. Crane ◽  
Angel J. Di Bilio ◽  
...  
Keyword(s):  
Monoclinic Space Group ◽  
Native Protein ◽  
Unit Cell Parameters ◽  
Space Group ◽  
X Ray ◽  
Cell Parameters ◽  
X Ray Crystallography ◽  
Peg 4000 ◽  
Dark Green ◽  
Atomic Coordinates

The crystal structure of Ru(2,2′-bipyridine)2(imidazole)(His83)azurin (RuAz) has been determined to 2.3 Å resolution by X-ray crystallography. The spectroscopic and thermodynamic properties of both the native protein and [Ru(2,2′-bipyridine)2(imidazole)2]2+ are maintained in the modified protein. Dark-green RuAz crystals grown from PEG 4000, LiNO3, CuCl2 and Tris buffer are monoclinic, belong to the space group C2 and have cell parameters a = 100.6, b = 35.4, c = 74.7 Å and β =  106.5°. In addition, [Ru(2,2′-bipyridine)2(imidazole)2]SO4·10H2O was synthesized, crystallized and structurally characterized by X-ray crystallography. Red–brown crystals of this complex are monoclinic, space group P21/n, unit-cell parameters a = 13.230 (2), b = 18.197 (4), c = 16.126 (4) Å, β = 108.65 (2)°. Stereochemical parameters for the refinement of Ru(2,2′-bipyridine)2(imidazole)(His83) were taken from the atomic coordinates of [Ru(2,2′-bipyridine)2(imidazole)2]2+. The structure of RuAz confirms that His83 is the only site of chemical modification and that the native azurin structure is not perturbed significantly by the ruthenium label.

scholarly journals Crystallization and X-ray diffraction studies of a two-domain laccase fromStreptomyces griseoflavus

2015 ◽  
Vol 71 (9) ◽  
pp. 1200-1204 ◽  
Author(s):  
Svetlana Tishchenko ◽  
Azat Gabdulkhakov ◽  
Liubov Trubitsina ◽  
Alexander Lisov ◽  
Marina Zakharova ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Molecular Weight ◽  
Phenolic Compounds ◽  
Synchrotron Radiation ◽  
Monoclinic Space Group ◽  
Asymmetric Unit ◽  
X Ray Diffraction ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters

Laccase (EC 1.10.3.2) is one of the most common copper-containing oxidases; it is found in many organisms and catalyzes the oxidation of primarily phenolic compounds by oxygen. Two-domain laccases have unusual thermostability, resistance to inhibitors and an alkaline optimum of activity. The causes of these properties in two-domain laccases are poorly understood. A recombinant two-domain laccase (SgfSL) was cloned from the genome ofStreptomyces griseoflavusAc-993, expressed inEscherichia coliand purified to homogeneity. The crystals of SgfSL belonged to the monoclinic space groupP21, with unit-cell parametersa= 74.64,b= 94.72,c= 117.40 Å, β = 90.672°, and diffraction data were collected to 2.0 Å resolution using a synchrotron-radiation source. Two functional trimers per asymmetric unit correspond to a Matthews coefficient of 1.99 Å3 Da−1according to the monomer molecular weight of 35.6 kDa.

scholarly journals Expression, high-pressure refolding, purification, crystallization and preliminary X-ray analysis of a novel single-strand-specific 3′–5′ exonuclease PhoExo I fromPyrococcus horikoshiiOT3

2014 ◽  
Vol 70 (8) ◽  
pp. 1076-1079 ◽  
Author(s):  
Ken-ichi Miyazono ◽  
Kanae Tsutsumi ◽  
Yoshizumi Ishino ◽  
Masaru Tanokura
Keyword(s):  
High Pressure ◽  
Inclusion Bodies ◽  
Single Strand ◽  
Diffusion Method ◽  
X Rays ◽  
Asymmetric Unit ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters ◽  
Reservoir Solution

PhoExo I is a single-strand-specific 3′–5′ exonuclease fromPyrococcus horikoshiiOT3 and is thought to be involved in a Thermococcales-specific DNA-repair pathway. The recombinant PhoExo I protein was produced as inclusion bodies inEscherichia colicells. Solubilization of the inclusion bodies was performed by the high-pressure refolding method and highly purified protein was subjected to crystallization by the sitting-drop vapour-diffusion method at 20°C. A crystal of PhoExo I was obtained in a reservoir solution consisting of 0.1 MTris–HCl pH 8.9, 27% PEG 6000 and diffracted X-rays to 1.52 Å resolution. The crystal of PhoExo I belonged to space groupH32, with unit-cell parametersa=b= 112.07,c= 202.28 Å. The crystal contained two PhoExo I molecules in the asymmetric unit.

scholarly journals Crystallographic analysis of NosA, which catalyzes terminal amide formation in the biosynthesis of nosiheptide

2015 ◽  
Vol 71 (8) ◽  
pp. 1033-1037 ◽  
Author(s):  
Yanting Wang ◽  
Shanshan Liu ◽  
Pengfei Yao ◽  
Yi Yu ◽  
Yan Zhang ◽  
...  
Keyword(s):  
Synchrotron Radiation ◽  
Diffraction Data ◽  
Crystallographic Analysis ◽  
Diffusion Method ◽  
Asymmetric Unit ◽  
Unit Cell Parameters ◽  
Solvent Content ◽  
Cell Parameters ◽  
Vapour Diffusion ◽  
Streptomyces Actuosus

Nosiheptide is a member of the thiopeptide family of antibiotics which demonstrates potent activities against various bacterial pathogens. The formation of its C-terminal amide is catalysed by NosA in an unusual strategy for maturating certain thiopeptides by processing precursor peptides featuring a serine extension. Here, a recombinant C-terminally truncated selenomethionine-derivatized NosA1–111variant fromStreptomyces actuosusconsisting of residues 1–111, named SeMet NosA1–111, was crystallized using the sitting-drop vapour-diffusion method. Diffraction data were collected to 2.40 Å resolution using synchrotron radiation. The crystals belonged to the primitive cubic space groupP4132, with unit-cell parametersa=b=c= 143.3 Å. Assuming the presence of three molecules in the asymmetric unit, the calculated Matthews coefficient was 3.94 Å3 Da−1and the corresponding solvent content was 40.3%.

scholarly journals Crystallization and preliminary X-ray diffraction analysis of the Csu pili CsuC–CsuA/B chaperone–major subunit pre-assembly complex fromAcinetobacter baumannii

2015 ◽  
Vol 71 (6) ◽  
pp. 770-774 ◽  
Author(s):  
Natalia Pakharukova ◽  
Minna Tuittila ◽  
Sari Paavilainen ◽  
Anton Zavialov
Keyword(s):  
Diffusion Method ◽  
X Ray Diffraction ◽  
Hanging Drop ◽  
Unit Cell Parameters ◽  
Gram Negative ◽  
X Ray ◽  
Cell Parameters ◽  
Abiotic Surfaces ◽  
Vapour Diffusion ◽  
Fimbrial Adhesins

The attachment of many Gram-negative pathogens to biotic and abiotic surfaces is mediated by fimbrial adhesins, which are assembledviathe classical, alternative and archaic chaperone–usher (CU) pathways. The archaic CU fimbrial adhesins have the widest phylogenetic distribution, yet very little is known about their structure and mechanism of assembly. To elucidate the biogenesis of archaic CU systems, structural analysis of the Csu fimbriae, which are used byAcinetobacter baumanniito form stable biofilms and cause nosocomial infection, was focused on. The major fimbriae subunit CsuA/B complexed with the CsuC chaperone was purified from the periplasm ofEscherichia colicells co-expressing CsuA/B and CsuC, and the complex was crystallized in PEG 3350 solution using the hanging-drop vapour-diffusion method. Selenomethionine-labelled CsuC–CsuA/B complex was purified and crystallized under the same conditions. The crystals diffracted to 2.40 Å resolution and belonged to the hexagonal space groupP6422, with unit-cell parametersa=b= 94.71,c = 187.05 Å, α = β = 90, γ = 120°. Initial phases were derived from a single anomalous diffraction (SAD) experiment using the selenomethionine derivative.

Refinement of the crystal structure of sieleckiite and revision of its symmetry

2017 ◽  
Vol 81 (4) ◽  
pp. 917-922
Author(s):  
Peter Elliott
Keyword(s):  
Crystal Structure ◽  
Synchrotron Radiation ◽  
Single Crystal ◽  
Diffraction Data ◽  
X Ray Diffraction ◽  
Aluminium Phosphate ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters ◽  
Phosphate Mineral

AbstractThe crystal structure of the copper aluminium phosphate mineral sieleckiite, Cu3Al4(PO4)2 (OH)12·2H2O, from the Mt Oxide copper mine, Queensland, Australia was solved from single-crystal X-ray diffraction data utilizing synchrotron radiation. Sieleckiite has monoclinic rather than triclinic symmetry as previously reported and is space group C2/m with unit-cell parameters a = 11.711(2), b = 6.9233(14), c = 9.828(2) Å, β = 92.88(3)°, V = 795.8(3) Å3and Z = 2. The crystal structure, which has been refined to R1 = 0.0456 on the basis of 1186 unique reflections with Fo > 4σF, is a framework of corner-, edge- and face- sharing Cu and Al octahedra and PO4 tetrahedra.

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