scholarly journals The quorum-quenching lactonase from Alicyclobacter acidoterrestris: purification, kinetic characterization, crystallization and crystallographic analysis

2017 ◽  
Vol 73 (8) ◽  
pp. 476-480 ◽  
Author(s):  
Celine Bergonzi ◽  
Michael Schwab ◽  
Eric Chabriere ◽  
Mikael Elias
Keyword(s):  
Quorum Quenching ◽  
Crystallographic Analysis ◽  
Gram Negative Bacteria ◽  
Kinetic Characterization ◽  
Chemical Signaling ◽  
X Ray Diffraction ◽  
Bacterial Communication ◽  
X Ray ◽  
Molecular Determinants ◽  
Engineering Projects

Lactonases comprise a class of enzymes that hydrolyze lactones, including acyl-homoserine lactones (AHLs); the latter are used as chemical signaling molecules by numerous Gram-negative bacteria. Lactonases have therefore been demonstrated to quench AHL-based bacterial communication. In particular, lactonases are capable of inhibiting bacterial behaviors that depend on these chemicals, such as the formation of biofilms or the expression of virulence factors. A novel representative from the metallo-β-lactamase superfamily, named AaL, was isolated from the thermoacidophilic bacterium Alicyclobacter acidoterrestris. Kinetic characterization proves AaL to be a proficient lactonase, with catalytic efficiencies (k cat/K m) against AHLs in the region of 105 M −1 s−1. AaL exhibits a very broad substrate specificity. Its structure is expected to reveal the molecular determinants for its substrate binding and specificity, as well as to provide grounds for future protein-engineering projects. Here, the expression, purification, characterization, crystallization and X-ray diffraction data collection of AaL at 1.65 Å resolution are reported.

scholarly journals The quorum-quenching lactonase fromGeobacillus caldoxylosilyticus: purification, characterization, crystallization and crystallographic analysis

2016 ◽  
Vol 72 (9) ◽  
pp. 681-686 ◽  
Author(s):  
Celine Bergonzi ◽  
Michael Schwab ◽  
Mikael Elias
Keyword(s):  
Catalytic Mechanism ◽  
Quorum Quenching ◽  
Thermophilic Bacterium ◽  
Thermostable Enzyme ◽  
Crystallographic Analysis ◽  
Chemical Signaling ◽  
X Ray Diffraction ◽  
Molecular Determinants ◽  
Interesting Candidate ◽  
Shed Light

Lactonases are enzymes that are capable of hydrolyzing various lactones such as aliphatic lactones or acyl-homoserine lactones (AHLs), with the latter being used as chemical signaling molecules by numerous Gram-negative bacteria. Lactonases therefore have the ability to quench the chemical communication, also known as quorum sensing, of numerous bacteria, and in particular to inhibit behaviors that are regulated by this system, such as the expression of virulence factors or the production of biofilms. A novel representative from the metallo-β-lactamase superfamily, dubbed GcL, was isolated from the thermophilic bacteriumGeobacillus caldoxylosilyticus. Because of its thermophilic origin, GcL may constitute an interesting candidate for the development of biocontrol agents. Here, we show that GcL is a thermostable enzyme with a half-life at 75°C of 152.5 ± 10 min. Remarkably, it is also shown that GcL is among the most active lactonases characterized to date, with catalytic efficiencies (kcat/Km) against AHLs of greater than 106 M−1 s−1. The structure of GcL is expected to shed light on the catalytic mechanism of the enzyme and the molecular determinants for the substrate specificity in this class of lactonases. Here, the expression, purification, characterization, crystallization and X-ray diffraction data collection to 1.6 Å resolution of GcL are reported.

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.

Synthesis and Characterization of Cadmium with Titanium Oxide (Cd-TiO2) Nanocomposite: Testing its Antibacterial Effect

2021 ◽  
Vol 12 (4) ◽  
pp. 2523-2529
Author(s):  
Daniel Sam N ◽  
Anish C I ◽  
Sabeena G ◽  
Rajaduraipandian S ◽  
Manobala ◽  
...  
Keyword(s):  
Sol Gel ◽  
Gram Negative Bacteria ◽  
X Ray Diffraction ◽  
Sem Images ◽  
Gram Negative ◽  
X Ray ◽  
Weight Percentage ◽  
Inhibition Concentration ◽  
Xrd Patterns

Sol gel methods were used for the study of the antimicrobial activity of Cd-TiO2 against gram-negative and positive bacteria. These Cd-TiO2 have been characterized by various optical and techniques. They have been exhibited by X-ray diffraction, scanning electron microscopy, ultraviolet spectroscopy, and infrared spectroscopy. The structures of the various XRD patterns indicate that the product has a structure. The particle size of Cd-TiO2 is 35nm. The SEM images confirm the spherical appearance of the sample. The energy X-ray spectra have been confirmed as well and then C, O, Ti, Cd, Pt element are present in Cd-TiO2. The weight percentage of Cadmium is 5.8%, Ti is 51.03%, C is 5.13% and O is 31.75% in Cd-TiO2. BET image shows that the major pore size distribution of Cd-TiO2 is ranged from 2.24 nm. The Cd-TiO2 that the antibacterial activity when tested against the pathogens only gram-negative bacteria such as Pseudomonas. The zone of minimum inhibition concentration was measured in a range of 20mm in 25μl and 30mm in 100μl.

X-ray diffraction has limited applicability in investigation of milk tampering

2019 ◽  
Vol 86 (3) ◽  
pp. 337-340
Author(s):  
Ana Paula Pavão Battaglini ◽  
Alexandre Urbano ◽  
Vanerli Beloti ◽  
Edson Antonio Rios ◽  
Juliana Ramos Pereira ◽  
...  
Keyword(s):  
Sodium Citrate ◽  
Freezing Point ◽  
Raw Milk ◽  
Physicochemical Analysis ◽  
Crystallographic Analysis ◽  
X Ray Diffraction ◽  
Water Addition ◽  
X Ray ◽  
Freeze Dried ◽  
Limited Applicability

AbstractThe aim of this work was to use X-ray diffraction to identify substances used for adulteration of raw milk and to determine if crystallographic analysis can detect extraneous substances in milk. Two unknown substances were sent anonymously by employers linked to the dairy chain, who claimed that they were added directly in milk prior to water addition by truck drivers. The samples were analyzed by X-ray diffraction and submitted to physicochemical analysis. The first substance was identified by X-ray diffraction as sodium citrate, complying with its physicochemical attributes, such as the powerful ability to decrease the freezing point. The second substance was identified by X-ray diffraction as sucrose and this result was also in agreement with its ability to increase the density, decrease the freezing point and finally, to be positive for sucrose in the resorcinol qualitative test. To evaluate if X-ray diffraction can detect extraneous substances already mixed in milk, fresh raw milk samples tampered with urea, sodium hydroxide, sodium citrate and sucrose were freeze dried and analyzed by X-ray diffraction, with no detection of any extraneous substances at any percentage. This is the first report of attempted diagnosis of extraneous substances in milk by X-ray diffraction. However, this technique can be useful only when applied to identify substances used for adulteration prior to its dilution in milk, since the amorphous nature of milk seems to be a limitation for the accurate detection of extraneous substances.

scholarly journals Crystallization and preliminary crystallographic analysis of manganese lipoxygenase

2014 ◽  
Vol 70 (4) ◽  
pp. 522-525 ◽  
Author(s):  
Anneli Wennman ◽  
Ernst H. Oliw ◽  
Saeid Karkehabadi
Keyword(s):  
Expression System ◽  
Crystallographic Analysis ◽  
Gaeumannomyces Graminis ◽  
X Ray Diffraction ◽  
X Ray ◽  
Cell Parameters ◽  
Apparent Homogeneity ◽  
Structural Differences ◽  
Position Specificity ◽  
Take All

Lipoxygenases constitute a family of nonhaem metal enzymes with catalytic iron or, occasionally, catalytic manganese. Lipoxygenases oxidize polyunsaturated fatty acids with position specificity and stereospecificity to hydroperoxides, which contribute to inflammation and the development of cancer. Little is known about the structural differences between lipoxygenases with Fe or Mn and the metal-selection mechanism. APichia pastorisexpression system was used for the production of the manganese lipoxygenase of the take-all fungus of wheat,Gaeumannomyces graminis. The active enzyme was treated with α-mannosidase, purified to apparent homogeneity and subjected to crystal screening and X-ray diffraction. The crystals diffracted to 2.6 Å resolution and belonged to space groupC2, with unit-cell parametersa= 226.6,b= 50.6,c= 177.92 Å, β = 91.70°.

scholarly journals Crystallization and preliminary X-ray crystallographic analysis of carboxyl-terminal region 4 of SigR fromStreptomyces coelicolorA3(2)

2014 ◽  
Vol 70 (6) ◽  
pp. 747-749
Author(s):  
Keon Young Kim ◽  
Sunmin Kim ◽  
Jeong Kuk Park ◽  
HyoJin Song ◽  
SangYoun Park
Keyword(s):  
Streptomyces Coelicolor ◽  
Crystallographic Analysis ◽  
Terminal Region ◽  
Full Length ◽  
Spectrometric Analysis ◽  
X Rays ◽  
X Ray Diffraction ◽  
X Ray ◽  
Cell Parameters ◽  
Carboxyl Terminal

Full-length SigR fromStreptomyces coelicolorA3(2) was overexpressed inEscherichia coli, purified and submitted to crystallization trials using either polyethylene glycol 3350 or 4000 as a precipitant. X-ray diffraction data were collected to 2.60 Å resolution under cryoconditions using synchrotron X-rays. The crystal packs in space groupP43212, with unit-cell parametersa=b= 42.14,c= 102.02 Å. According to the Matthews coefficient, the crystal asymmetric unit cannot contain the full-length protein. Molecular replacement with the known structures of region 2 and region 4 as independent search models indicates that the crystal contains only the −35 element-binding carboxyl-terminal region 4 of full-length SigR. Mass-spectrometric analysis of the harvested crystal confirms this, suggesting a crystal volume per protein weight (VM) of 2.24 Å3 Da−1and 45.1% solvent content.

scholarly journals Cloning, expression, purification, crystallization and preliminary crystallographic analysis of 5-aminolaevulinic acid dehydratase fromBacillus subtilis

2010 ◽  
Vol 66 (9) ◽  
pp. 1053-1055 ◽  
Author(s):  
Qianda Lu ◽  
Jinming Ma ◽  
Hui Rong ◽  
Jun Fan ◽  
Ye Yuan ◽  
...  
Keyword(s):  
Crystallographic Analysis ◽  
Diffusion Method ◽  
X Ray Diffraction ◽  
Gene Encoding ◽  
Data Set ◽  
X Ray ◽  
Aminolaevulinic Acid ◽  
Acid Dehydratase ◽  
Strain Bl21 ◽  
Aminolaevulinic Acid Dehydratase

5-Aminolaevulinic acid dehydratase (ALAD), a crucial enzyme in the biosynthesis of tetrapyrrole, catalyses the condensation of two 5-aminolaevulinic acid (ALA) molecules to form porphobilinogen (PBG). The gene encoding ALAD was amplified from genomic DNA ofBacillus subtilisand the protein was overexpressed inEscherichia colistrain BL21 (DE3). The protein was purified and crystallized with an additional MGSSHHHHHHSSGLVPRGSH– tag at the N-terminus of the target protein. Diffraction-quality single crystals were obtained by the hanging-drop vapour-diffusion method. An X-ray diffraction data set was collected at a resolution of 2.7 Å.

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.

Mannich Reaction Product of Quinolin-8-ol (Oxine) and its Antibacterial Activity

1997 ◽  
Vol 50 (8) ◽  
pp. 861 ◽  
Author(s):  
Shaheen Faizi ◽  
Bina Shaheen Siddiqui ◽  
Rubeena Saleem ◽  
Farzana Akhtar ◽  
Khursheed Ali Khan ◽  
...  
Keyword(s):  
Antibacterial Activity ◽  
Single Crystal ◽  
Mannich Reaction ◽  
Gram Negative Bacteria ◽  
X Ray Diffraction ◽  
Gram Positive ◽  
Gram Negative ◽  
Antimicrobial Screening ◽  
X Ray

Quinolin-8-ol (1) on reaction with 2,6-dichloroaniline and m-nitrobenzaldehyde afforded 7-(α-hydroxy- m-nitrobenzyl)quinolin-8-ol (2), which is an unusual Mannich reaction product. The structure of (2) was determined by spectroscopic and single-crystal X-ray diffraction studies. Preliminary antimicrobial screening showed that (1) and (2) have activity against a large number of Gram-positive and Gram-negative bacteria.

Sign in / Sign up

Export Citation Format

Share Document