A High-Homology Region Provides the Possibility of Detecting β-Barrel Pore-Forming Toxins from Various Bacterial Species.

The pathogenicity of many bacteria, including and , depends on pore-forming toxins (PFTs), which cause the lysis of host cells by forming pores in the membranes of eukaryotic cells. Bioinformatic analysis revealed a region homologous to the Lys171-Gly250 sequence in hemolysin II (HlyII) from in over 600 PFTs, which we designated as a "homologous peptide". Three β-barrel PFTs were used for a detailed comparative analysis.

Genomic analysis and assessment of pathogenic (toxicogenic) potential of Staphylococcus haemolyticus and Bacillus paranthracis consortia isolated from bovine mastitis in Russia.

Three stable microbial consortia, each composed of Bacillus paranthracis and Staphylococcus haemolyticus strains, were isolated from milk of cows diagnosed with mastitis in three geographically remote regions of Russia. The composition of these consortia remained stable following multiple passages on culture media. Apparently, this stability is due to the structure of the microbial biofilms formed by the communities.

The Effects of Zirconium and Yttrium Addition on the Microstructure and Hardness of AlCuMgMn Alloy when Applying In Situ Heating during the Laser Melting Process.

This paper studies the effect of the laser melting process (LMP) on the microstructure and hardness of a new modified AlCuMgMn alloy with zirconium (Zr) and Yttrium (Y) elements. Homogenized (480 °C/8 h) alloys were laser-surface-treated at room temperature and a heating platform with in situ heating conditions was used in order to control the formed microstructure by decreasing the solidification rate in the laser-melted zone (LMZ). Modifying the AlCuMgMn alloy with 0.

Region Met225 to Ile412 of Hemolysin II Is Capable to Agglutinate Red Blood Cells.

Hemolysin II (HlyII) is one of the virulence factors of the opportunistic bacterium belonging to the group of β-pore-forming toxins. This work created a genetic construct encoding a large C-terminal fragment of the toxin (HlyIILCTD, M225-I412 according to the numbering of amino acid residues in HlyII). A soluble form of HlyIILCTD was obtained using the SlyD chaperone protein.

The C-terminal domain of Bacillus cereus hemolysin II oligomerizes by itself in the presence of cell membranes to form ion channels.

Bacillus cereus hemolysin II, a pore-forming β-barrel toxin (HlyII), has a C-terminal extension of 94 amino acid residues, designated as the C-terminal domain of HlyII (HlyIICTD). HlyIICTD is capable of forming oligomers in aqueous solutions. Oligomerization of HlyIICTD significantly increased in the presence of erythrocytes and liposomes.

Influence of Adding Modifying Elements and Homogenization Annealing on Laser Melting Process of the Modified AlZnMgCu with 4%Si Alloys.

AlZnMgCu, the high-strength aluminum alloy, is unsuitable for laser melting applications due to its high hot cracking sensitivity and large solidification temperature range. Adapting this alloy for laser melting processing is a high-demand research issue for extending its use. Thus, this paper investigates the effect of adding 4%Si, 4%Si-Sc+Zr, 4%Si-Ti+B, and homogenization annealing on the laser melting process (LMP) of AlZnMgCu alloy.

A simple method to purify recombinant HCV core protein expressed in Pichia pastoris for obtaining virus-like particles and producing monoclonal antibodies.

In this study, we describe an optimized method of obtaining virus-like particles (VLPs) of the recombinant hepatitis C virus (HCV) core protein (HCcAg) expressed in yeast cells (Pichia pastoris), which can be used for the construction of diagnostic test systems and vaccine engineering. The described simplified procedure was developed to enable in vitro self-assembly of HCcAg molecules into VLPs during protein purification. In brief, the HCcAg protein was precipitated from yeast cell lysates with ammonium sulfate and renatured by gel filtration on Sephadex G-25 under reducing conditions.

Monoclonal Antibody HlyIIC‑15 to C-End Domain HlyII Interacts with the Trombin Recognition Site.

Among the panel of monoclonal antibodies to the recombinant protein HlyIICTD an antibody was found capable of forming an immune complex with a thrombin recognition region, the amino acid sequence of which is located inside the recombinant HlyIICTD. Localization of the epitope was carried out using peptide phage display methods, as well as enzyme immunoassay and immunoblotting for interaction with recombinant proteins, either containing or not containing individual components HlyIICTD. The identified epitope is located in the region of the thrombin site and retains the ability to interact with the antibody after the proteolyotic attack of the protein by thrombin.

A Monoclonal Antibody against the C-Terminal Domain of Hemolysin II Inhibits HlyII Cytolytic Activity.

is the fourth most common cause of foodborne illnesses that produces a variety of pore-forming proteins as the main pathogenic factors. hemolysin II (HlyII), belonging to pore-forming β-barrel toxins, has a C-terminal extension of 94 amino acid residues designated as HlyIICTD. An analysis of a panel of monoclonal antibodies to the recombinant HlyIICTD protein revealed the ability of the antibody HlyIIC-20 to inhibit HlyII hemolysis.

Evaluation of the Microstructure and Mechanical Properties of a New Modified Cast and Laser-Melted AA7075 Alloy.

The mechanical properties and microstructure of as-cast and homogenized AA7075 were investigated. This alloy was modified by adding transition elements 0.3%Sc + 0.

Desktop Fabrication of Strong Poly (Lactic Acid) Parts: FFF Process Parameters Tuning.

The current study aims to evaluate the possibilities to increase part strength by optimizing the Fused Filament Fabrication (FFF) process parameters. Five different CAD models of parts with the same coupling dimensions but of different shape inherited from a recent study were converted into test samples with Ultimaker 2 3D printer. The main measure of success was the sample strength, defined as the load at which the first crack in the stressed area of the part appeared.

An alternative approach to study the enzymatic specificities of the CfrBI restriction-modification system.

Restriction-modification systems (RMS) are the main gene-engineering tools and a suitable model to study the molecular mechanisms of catalysis and DNA-protein interactions. Research into the catalytic properties of these enzymes, determination of hydrolysis and DNA-methylation sites remain topical. In our previous work we have cloned and sequenced the CfrBI restriction-modification system (strain , which recognizes the nucleotide sequence 5'-CCWWGG-3'.

Design and Fabrication of Strong Parts from Poly (Lactic Acid) with a Desktop 3D Printer: A Case with Interrupted Shell.

The ability to form closed cavities inside the part printed is an important feature of Fused Filament Fabrication technology. A typical part consists of a dense shell bearing the primary load, filled with low-density plastic scaffold (infill). Such a constitution of the part provides in most cases appropriate strength and low weight.

Strength of PLA Components Fabricated with Fused Deposition Technology Using a Desktop 3D Printer as a Function of Geometrical Parameters of the Process.

The current paper studies the influence of geometrical parameters of the fused deposition modeling (FDM)-fused filament fabrication (FFF) 3D printing process on printed part strength for open source desktop 3D printers and the most popular material used for that purpose-i.e., polylactic acid (PLA).

Hepatic and Aortic Arch Expression and Serum Levels of Syndecan-1 in ApoE Mice.

Background: Heparan sulfate proteoglycan (HSPG) syndecan-1 (Sdc1) acts as a receptor for triglyceride-rich lipoproteins (TRLs), growth factors, chemokines and enzymes. Due to the disordered structure, its function is as diverse as its ligands. In this paper, we have analyzed hepatic and aortic arch expression of Sdc1 in ApoE mice and examined their association with biochemical changes in plasma during the atheroma formation.

Protein NCRII-18: the role of gene fusion in the molecular evolution of restriction endonucleases.

This work first constructed the fusion protein NCRII-18 by fusing the restriction endonuclease Ecl18kI gene and part of the gene coding for the N-terminal domain of the endonuclease EcoRII. The fusion of the EcoRII N-terminal domain leads to a change in the properties of the recombinant protein. Unlike Ecl18kI, which made the basis of NCRII-18, the fusion protein predominantly recognizes the CCWGG sites, having lost the capability of interacting with the CCSGG sites.

Xylanases of Cellulomonas flavigena: expression, biochemical characterization, and biotechnological potential.

Four xylanases of Cellulomonas flavigena were cloned, expressed in Escherichia coli and purified. Three enzymes (CFXyl1, CFXyl2, and CFXyl4) were from the GH10 family, while CFXyl3 was from the GH11 family. The enzymes possessed moderate temperature stability and a neutral pH optimum.

[Effect of Bacillus cereus hemolysin II on hepatocyte cells].

We investigated the efficiency of increasing the permeability (permeabilization) of cell membranes in primary liver cells by Bacillus cereus hemolysin II. An assessment of the degree of permeabilization was car ried out by measuring the fluorescence intensity of various low molecular weight dyes, which enter through pores into hepatocyte cells cultivated with hemolysin. We uncovered a high efficacy of hemolysin HlyII action on hepatocyte cell walls, which exceeded the effect of nonionic detergent, digitonin, which is commonly employed for pore formation in various cell membranes.

Preparation of mononucleosomal templates for analysis of transcription with RNA polymerase using spFRET.

Single positioned nucleosomes have been extensively employed as simple model experimental systems for analysis of various intranuclear processes. Here we describe an experimental system containing positioned mononucleosomes allowing transcription by various RNA polymerases. Each DNA template contains a pair of fluorescent labels (Cy3 and Cy5) allowing measuring relative distances between the neighboring coils of nucleosomal DNA using Forster resonance energy transfer (FRET).

[Purification of recombinant Bacillus cereus ResD-ResE proteins expressed in Escherichia coli strains].

Recombinant E. coli strains expressing the Bacillus cereus ATCC 14579T resD and resEgenes fused with the ubiquitin gene were constructed, and purification of the ResD and ResE proteins was performed. The approach used in the study allowed us to increase the protein yield of the electrophoretic homogeneous ResD andResE proteins without denaturation steps up to 150 mg per gram of wet cell weight.

Role of structural changes induced in biological membranes by hydrolysable tannins from sumac leaves (Rhus typhina L.) in their antihemolytic and antibacterial effects.

In this study, we found that the sumac tannins (Rhus typhina L.) exert to a various extent antihemolytic effects and antibacterial activity against Bacillus cereus and Pseudomonas aeruginosa depending on structural specificity of bacteria and different mechanisms of their toxic action. The sumac tannins exert the most expressed activity against B.

Recombinant xylanase from Streptomyces coelicolor Ac-738: characterization and the effect on xylan-containing products.

A xylanase gene was isolated from the genomic DNA of Streptomyces coelicolor Ac-738. The 723-bp full-length gene encoded a 241-amino acid peptide consisting of a 49-residue putative TAT signal peptide and a glycoside hydrolase family-11 domain. The mature enzyme called XSC738 was expressed in Escherichia coli M15[pREP4].

Binding of DNA methyltransferase M.Ecl18kI [corrected] to operator-promoter region decreases its methylating activity.

The type II bifunctional DNA methyltransferase (MTase) Ecl18 that is able to control transcription of its own gene was studied kinetically. Based on initial velocity dependences from S-adenosyl-L-methionine (AdoMet) and target DNA and substrate preincubation assays, it is proposed that the enzyme apparently works by a rapid equilibrium ordered bi-bi mechanism with DNA binding first. By measuring the enzyme activity depending on DNA and AdoMet at different fixed concentrations of the operator sequence oligonucleotide, it was found that its binding has noncompetitive inhibitory effect on Ecl18 MTase activity.