Isoelectric Point Values Research Articles

Effects of detoxification process on toxicity and foreign protein of tetanus toxoid and diphtheria toxoid

Formaldehyde detoxification is a process for converting tetanus toxin (TT) and diphtheria toxin (DT) into tetanus toxoid (TTd) and diphtheria toxoid (DTd), respectively. The mechanism of this detoxification process has been investigated by several previous studies based on lab-scale toxoids. To obtain greater insights of the effects induced by formaldehyde, industrial TTd and DTd batches obtained from different detoxification processes were studied in this work. Using liquid chromatography-mass spectrometry (LC-MS), 15 and 20 repeatable formaldehyde-induced modification sites of TTd and DTd were identified, respectively. Toxoid which had a higher formaldehyde-induced modification rate observed by LC-MS, also had larger bands on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Aggregates which were observed on size exclusion chromatogram (SEC) were confirmed by SDS-PAGE and LC-MS. Formaldehyde detoxification also led to a decrease of isoelectric point (pI) values and an increase of retention on weak anion exchange (WAX) column. Specific toxicity tests were conducted to evaluate toxicity of the TTd and DTd samples obtained with different detoxification conditions. Results from the specific toxicity tests showed that all toxoids used in this study were qualified, including toxoids obtained from mild and drastic detoxification conditions. However, obtained from mild detoxification conditions had less aggregates and may lead to a higher degree of glycosylation in conjugate vaccines than the ones obtained from drastic detoxification conditions. Thus, we suggest that mild detoxification conditions should be used to obtain TTd and DTd. Furthermore, as well as studying the formaldehyde-induced modifications and toxicity in TTd and DTd, the effects of the detoxification process on foreign proteins were also investigated. An increase in foreign proteins were observed in the aggregate than in the monomer of the toxoids. Additionally, some foreign proteins in the monomer of the toxins transferred to the aggregate of toxoids due to the formation of cross-linking. To eliminate the risk of cross-linking foreign proteins to toxoids in vaccination programs, a purification process is necessary before the detoxification process and/or the use of toxoids in vaccines.

SHTXTHHly, an extracellular secretion platform for the preparation of bioactive peptides and proteins in Escherichia coli

BackgroundIn previous work, we developed an E. coli extracellular secretion platform XTHHly based on the hemolysin A secretion system. It can produce bioactive peptides with simple purification procedures. However, the wider application of this platform is limited by poor secretion efficiency.ResultsIn this study, we first discovered a positive correlation between the isoelectric point (pI) value of the target protein and the secretion level of the XTHHly system. Given the extremely high secretion level of S tag, we fused it at the N-terminus and created a novel SHTXTHHly system. The SHTXTHHly system significantly increased the secretion levels of antimicrobial peptides (PEW300, LL37, and Aurein 1.2) with full bioactivities, suggesting its excellent capacity for secretory production of bioactive peptides. Furthermore, RGDS, IL-15, and alcohol dehydrogenase were successfully secreted, and their bioactivities were largely maintained in the fusion proteins, indicating the potential applications of the novel system for the rapid determination of protein bioactivities. Finally, using the SHTXTHHly system, we produced the monomeric Fc, which showed a high affinity for Fcγ Receptor I and mediated the antibody-dependent immunological effects of immune cells, demonstrating its potential applications in immunotherapies.ConclusionsThe SHTXTHHly system described here facilitates the secretory production of various types of proteins in E. coli. In comparison to previously reported expression systems, our work enlightens an efficient and cost-effective way to evaluate the bioactivities of target proteins or produce them.Graphical abstract

Polyvinylidene fluoride/boehmite nanocomposite membrane for effective removal of arsenate ion from water

Groundwater arsenic contamination is a serious threat that has affected the health of 130 million people globally. Long-term exposure to arsenic leads to arsenic poisoning, i.e., arsenicosis. Therefore, the development of new technology to reject arsenic from water has been increased. Polyvinylidene fluoride (PVDF) polymer possesses outstanding properties mainly as thermal stability, chemical and mechanical resistance. Boehmite, a polymorphic aluminium oxide, is an essential industrial mineral used as an adsorbent due to its ability to retain high surface area. It has a positive charge, and its surface renders it easy for compounds with negative charges to be attached by electrostatic attraction. Also, the nanoparticles have a high isoelectric value. High adsorption is expected on a higher isoelectric point value. PVDF/boehmite composite membrane was prepared using solvent casting method at room temperature. The nanoparticles were added in Al/As (w/w) ratio of 0:1, 50:1, 200:1 and 500:1. Al/As ratio of 200:1 has shown the best result in batch process, and in static adsorption study, 65% rejection of arsenate ion was achieved from 100 μg/L arsenate solution in 1 h. Flux was calculated in a dead-end filtration setup at 1 bar trans-membrane pressure. The water flux of the membranes was in the range of 62 to 270 L/m2h.

Bioinformatic characterization of a triacylglycerol lipase produced by Aspergillus flavus isolated from the decaying seed of Cucumeropsis mannii

Lipases are enzymes of industrial importance responsible for the hydrolysis of ester bonds of triglycerides. A lipolytic fungus was isolated and subsequently identified based on the ITS sequence analysis as putative Aspergillus flavus with accession number LC424503. The gene coding for extracellular triacylglycerol lipase was isolated from Aspergillus flavus species, sequenced, and characterised using bioinformatics tools. An open reading frame of 420 amino acid sequence was obtained and designated as Aspergillus flavus lipase (AFL) sequence. Alignment of the amino acid sequence with other lipases revealed the presence GHSLG sequence which is the lipase consensus sequence Gly-X1-Ser-X2-Gly indicating that it a classical lipase. A catalytic active site lid domain composed of TYITDTIIDLS amino acids sequence was also revealed. This lid protects the active site, control the catalytic activity and substrate selectivity in lipases. The 3-Dimensional structural model shared 34.08% sequence identity with a lipase from Yarrowia lipolytica covering 272 amino acid residues of the template model. A search of the lipase engineering database using AFL sequence revealed that it belongs to the class GX-lipase, superfamily abH23 and homologous family abH23.02, molecular weight and isoelectric point values of 46.95 KDa and 5.7, respectively. N-glycosylation sites were predicted at residues 164, 236 and 333, with potentials of 0.7250, 0.7037 and 0.7048, respectively. O-glycosylation sites were predicted at residues 355, 358, 360 and 366. A signal sequence of 37 amino acids was revealed at the N-terminal of the polypeptide. This is a short peptide sequence that marks a protein for transport across the cell membrane and indicates that AFL is an extracellular lipase. The findings on the structural and molecular properties of Aspergillus flavus lipase in this work will be crucial in future studies aiming at engineering the enzyme for biotechnology applications. Communicated by Ramaswamy H. Sarma

The Filtration of 2D Electrophoresis Data during Creation of a Learning Set for Prediction of the Value of the Isoelectric Point of Proteins

A number of simple filters formulated from general considerations that take into account the peculiarities of the experiments as well as results obtained in 2D electrophoresis experiments are considered. These filters can be used for automated dataset formation and verification of learning of system for predicting protein isoelectric point values. These include: (i) filtering obvious errors introduced during initial database formation; (ii) selection of a known plausible range of values; (iii) selection of a single variant among various proteoforms; (iv) selection within a preset value of electrophoretic shift deviation, etc. Using a dataset combining data from 8 maps of Homo sapiens, Mus musculus, and Rattus norvegicus, the application of this set of filters improved the R2 value of predictions from 0.44 to 0.67.

PKPDB: a protein data bank extension database of pKa and pI theoretical values.

pKa values of ionizable residues and isoelectric points of proteins provide valuable local and global insights about their structure and function. These properties can be estimated with reasonably good accuracy using Poisson-Boltzmann and Monte Carlo calculations at a considerable computational cost (from some minutes to several hours). pKPDB is a database of over 12 M theoretical pKa values calculated over 120k protein structures deposited in the Protein Data Bank. By providing precomputed pKa and pI values, users can retrieve results instantaneously for their protein(s) of interest while also saving countless hours and resources that would be spent on repeated calculations. Furthermore, there is an ever-growing imbalance between experimental pKa and pI values and the number of resolved structures. This database will complement the experimental and computational data already available and can also provide crucial information regarding buried residues that are under-represented in experimental measurements. Gzipped csv files containing p Ka and isoelectric point values can be downloaded from https://pypka.org/pKPDB. To query a single PDB code please use the PypKa free server at https://pypka.org. The pKPDB source code can be found at https://github.com/mms-fcul/pKPDB. Supplementary data are available at Bioinformatics online.

Quercetin particles with lower inhibitory activity for α-glycosidase and negligible effects on blood clotting

Poly(quercetin) (p(QR)) particles was synthesis by using poly(ethylene glycol) diglycidyl ether (PEGGE) crosslinker in a single step via microemulsion system. The morphological, size and functional analysis of the prepared particles were carried by optic microscope, scanning electron microscope (SEM), dynamic light scattering (DLS) measurements and FT-IR spectroscopy. P(QR) particles were found to be in spherical shape with 372±9 nm size range by means of SEM images and DLS measurements. The zeta potential measurements, performed at different pH conditions and potentiometric titration of p(QR) particles were revealed that the isoelectric point and pKa values of as around pH 2.5 and 2.3, respectively. Ferric reducing antioxidant power (FRAP) was determined for QR and p(QR) particles at pH 3.6 and found as 9.4 and 0.43 µg reduced Fe(ll). The effects of QR and p(QR) particles on alfa-glycosidase enzyme activity at pH 6.9 were investigated and found that QR molecules and p(QR) particles were capable of inhibiting the α-glycosidase enzyme 89.3% and 24.7% respectively. The fluorescence spectroscopy of QR and p(QR) in fibrinogen showed that p(QR) particles do not induce clothing of blood.

Interactions between Phosphatidylcholine and Kaempferol or Myristicin: Langmuir Monolayers and Microelectrophoretic Studies.

Flavonoid compounds are known for their antibacterial, anti-inflammatory, and anticancer properties. Therefore, they can influence membrane properties that interest us, modifying both their structure and functions. We used kaempferol (K) and myricetin (M) as representatives of this group. We investigated the influence of the abovementioned compounds on model cell membranes’ properties (i.e., Langmuir monolayers and liposomes). The basic research methods used in these studies were the Langmuir method with Brewster angle microscopy and microelectrophoresis. The π–A isotherms were registered for the pure components and mixtures of these compounds with phosphatidylcholine (PC) in appropriate volume ratios. Using mathematical equations, we established that kaempferol, myricetin, and the lipids formed complexes at 1:1 ratios. We derived the parameters characterizing the formed complexes, i.e., the surfaces occupied by the complexes and the stability constants of the formed complexes. Using the microelectrophoretic method, we determined the dependence of the lipid membranes’ surface charge density as a function of the pH (in the range of 2 to 10) of the electrolyte solution. The presented results indicate that the PC membrane’s modification with kaempferol or myricetin affected changes in the surface charge density and isoelectric point values.

IPC 2.0: prediction of isoelectric point and pKa dissociation constants.

The isoelectric point is the pH at which a particular molecule is electrically neutral due to the equilibrium of positive and negative charges. In proteins and peptides, this depends on the dissociation constant (pKa) of charged groups of seven amino acids and NH+ and COO− groups at polypeptide termini. Information regarding isoelectric point and pKa is extensively used in two-dimensional gel electrophoresis (2D-PAGE), capillary isoelectric focusing (cIEF), crystallisation, and mass spectrometry. Therefore, there is a strong need for the in silico prediction of isoelectric point and pKa values. In this paper, I present Isoelectric Point Calculator 2.0 (IPC 2.0), a web server for the prediction of isoelectric points and pKa values using a mixture of deep learning and support vector regression models. The prediction accuracy (RMSD) of IPC 2.0 for proteins and peptides outperforms previous algorithms: 0.848 versus 0.868 and 0.222 versus 0.405, respectively. Moreover, the IPC 2.0 prediction of pKa using sequence information alone was better than the prediction from structure-based methods (0.576 versus 0.826) and a few folds faster. The IPC 2.0 webserver is freely available at www.ipc2-isoelectric-point.org

Genome-wide identification and expression pattern analysis of the ribonuclease T2 family in Eucommia ulmoides

The 2′,3′-cycling ribonuclease (RNase) genes are catalysts of RNA cleavage and include the RNase T2 gene family. RNase T2 genes perform important roles in plants and have been conserved in the genome of eukaryotic organisms. In this study we identified 21 EURNS genes in Eucommia ulmoides Oliver (E. ulmoides) and analyzed their structure, chromosomal location, phylogenetic tree, gene duplication, stress-related cis-elements, and expression patterns in different tissues. The length of 21 predicted EURNS proteins ranged from 143 to 374 amino acids (aa), their molecular weight (MW) ranged from 16.21 to 42.38 kDa, and their isoelectric point (PI) value ranged from 5.08 to 9.09. Two classifications (class I and class III) were obtained from the conserved domains analysis and phylogenetic tree. EURNS proteins contained a total of 15 motifs. Motif 1, motif 2, motif 3, and motif 7 were distributed in multiple sequences and were similar to the conserved domain of RNase T2. EURNS genes with similar structure and the predicted EURNS proteins with conserved motif compositions are in the same group in the phylogenetic tree. The results of RT-PCR and transcription data showed that EURNS genes have tissue-specific expression and exhibited obvious trends in different developmental stages. Gene duplication analysis results indicated that segment duplication may be the dominant duplication mode in this gene family. This study provides a theoretical basis for research on the RNase T2 gene family and lays a foundation for the further study of EURNS genes.

The bioinformatics of projected acetyl-CoA carboxylase genes in Hevea brasiliensis

This study clarified the variation and role of physical and chemical characteristics of the H. brasiliensis projected acetyl-CoA carboxylase genes. The detectable similarity and joining projected acetyl-CoA carboxylase genes of H. brasiliensis in the dendrogram may be the outcome of tropical environmental. Six projected acetyl-CoA carboxylase genes from H. Brasiliensis collected from NCBI. The NCBI locus number of the protein sequence utilized in this study was determined. The physical and chemical properties of acetyl-CoA carboxylase from H.brasiliensis. The factors were calculated the designate number of protein, number of amino acids, molecular weight, theoretical isoelectric point values, the total number of atoms, extinction coefficients, estimated half-life, instability index, aliphatic index and grand average of hydropathicity.

Rapid and Efficient Enrichment of Snake Venoms from Human Plasma Using a Strong Cation Exchange Tip Column to Improve Snakebite Diagnosis.

Snake envenomation is a serious public health issue in many tropical and subtropical countries. Accurate diagnosis and immediate antivenom treatment are critical for effective management. However, the venom concentration in the victims’ plasma is usually low, representing one of the bottlenecks in developing clinically applicable assays for venom detection and snakebite diagnosis. In this study, we attempted to develop a simple method for rapid enrichment of venom proteins from human plasma to facilitate detection. Our experiments showed that several major protein components of both Naja atra (N. atra) and Bungarus multicinctus (B. multicinctus) venoms have higher isoelectric point (pI) values relative to high-abundance human plasma proteins and could be separated via strong cation exchange–high-performance liquid chromatography (SCX-HPLC). Based on this principle, we developed an SCX tip column-based protocol for rapid enrichment of N. atra and B. multicinctus venom proteins from human plasma. Application of liquid chromatography-tandem mass spectrometry (LC-MS/MS) led to the identification of cytotoxin and beta-bungarotoxin as the major proteins enriched by the SCX tip column in each venom sample. The entire process of venom enrichment could be completed within 10–15 min. Combination of this method with our previously developed lateral flow strip assays (rapid test) significantly enhanced the sensitivity of the rapid test, mainly via depletion of the plasma protein background, as well as increase in venom protein concentration. Notably, the SCX tip column-based enrichment method has the potential to efficiently enrich other Elapidae snake venoms containing proteins with higher pI values, thereby facilitating venom detection with other assays. This simple and rapid sample preparation method should aid in improving the clinical utility of diagnostic assays for snakebite.

Structural-Genetic Characterization Of Novel Butaryl co-A Dehydrogenase and Proposition of Butanol Biosynthesis Pathway in Pusillimonas ginsengisoli SBSA.

Despite extensive use in the biofuel industry, only butyryl co-A dehydrogenase enzymes from the Clostridia group have undergone extensive structural and genetic characterization.The present study, portrays the characterization of structural, functional and phylogenetic properties of butyryl co-A dehydrogenase identified within the genome of Pusillimonas ginsengisoli SBSA. In silico characterization, homology modelling and docking data indicates that this protein is a homo-tetramer and 388 amino acid residue long, rich in alanine and leucine residue; having molecular weight of 42347.69 dalton. Its isoelectric point value is 5.78; indicate its neutral nature while 38.38 instability index value indicate its stable nature. Its thermostable nature evidenced by its high aliphatic index (93.14); makes its suitable for industry-based use. The secondary structure prediction analysis of butyryl co-A dehydrogenase unveiled that the proteins has secondary arrangements of 54% α-helix, 13% β-stand and 5% disordered conformation. However, phylogenetic analysis clearly indicates that probably horizontal gene transfer is the primary mechanism of spreading of this gene in this organism. Notably, multiple sequence alignment study of phylogenetically diverse butyryl co-A dehydrogenase sequence highlighted the presence of conserved amino acid residues i.e. YXV/LGXKXWXS/T. Physicochemical characterization of other relevant proteins involved in butanol metabolism of SBSA also has been carried out. However, metabolic construction of functional butanol biosynthesis pathway in SBSA, enlightened its cost-effective potential use in biofuel industry as an alternate to Clostridia system.

Fabrication, tuning and performance analysis of polyacrylonitrile (PAN)-derived microfiltration membranes for bacteria removal from drinking water

Removal of bacterial contaminations from water using advanced technologies is one of the essential steps in improving human health. The present study’s aim was to develop high performance microfiltration membranes from polyacrylonitrile (PAN) for bacteria removal from drinking water. The characteristics and performance of membranes were tuned through exploring the variation of prominent fabrication and operating parameters. The findings reveal that increasing PAN concentration in dope and addition of citric acid were successful in tailoring membrane microstructure. Bacteria rejection in modified membranes improved by exhibiting high log removal values (LRV) ranging from 3.92 (99.87%) to 5.57 (99.99%) while permeate fluxes were in the range of 35.83–58.62 L·m−2·h−1. The trends are explained by taking into account the structural characteristics of bacterial strains. Exploring the effect of operating parameters on the performance of membranes revealed that increase in feed concentration from 103 to 107 cfu·ml−1 improved membrane rejection. The largest rejection (5.57) was observed toward Staphylococcus aureus at feed concentration of 107 cfu·ml−1. Similarly, rejection improved upon reducing operating pressure from 3.5 to 1.5 bar. Also, shifting of feed pH to 7.4 and 9.4 enhanced membrane rejection to as high as 4.60 and 5.66 toward E. coli and Staphylococcus aureus, respectively. This was attributed to the zeta potential and isoelectric point values of the membranes and involved strains. Overall, the findings revealed that the developed PAN membranes are more effective in removal of gram-positive strains and their rejection is strongly dependent on the peptidoglycan layer of strains.

Identification and Analysis of <I>PGM</i> and <I>SUS</i> Gene Families in Leymus Chinensis Seed Transcriptome

Leymus chinensis ( Leymus chinensis  (Trin.) Tzvel.) is a good perennial forage with good palatability, high yield and high feeding value. In addition, because of its strong resistance, Leymus chinensis can improve the disadvantaged environment such as poor land, and it is an important species to protect grassland ecology in China. Therefore, in order to promote the germination of leymus chinensis seeds and explore the mechanism of Germination, this study used sterile water treatment of leymus chinensis seeds as control, exogenous GA3 as treatment conditions. Based on the analysis of pre-transcriptome data, it was determined that  PGM  and  SUS  genes in sucrose and starch metabolic pathways play a significant role in seed Germination. Members identification, conserved domains, and evolutionary trees were analyzed for both gene families using the bioinformatics method. The expression pattern of  PGM  and  SUS  genes were analyzed with FPKM value. The results showed that 7 members were selected from each of the the  PGM  and  SUS  families based on the data of Leymus Chinensis seeds transcriptome. The molecular weight of  PGM  genes ranged from 15 471.3 to 67 912.9 Da, and the isoelectric point value ranged from 4.45 to 6.31, showing weak acidity.  PGM  family in Leymus Chinensis seeds are hydrophilic stable proteins. The molecular weight of  SUS  genes ranged from 30 421.5 to 110 262.9 Da. The isoelectric point of  SUS  genes were neutral except  LcSUS1,5  and  7 , while the other four genes were weak acidic.  LcSUS4 , 6  were stable hydrophilic proteins, the rest were unstable hydrophilic proteins. The  PGM and  SUS  gene families in Leymus chinensis seeds play an enzyme role in promoting sucrose and starch synthesis in sucrose and starch metabolism pathways. The expression pattern analysis of Leymus chinensis seeds showed that the genes of  PGM  and  SUS  gene families in GA3-treated Leymus chinensis seeds promoted the germination of Leymus chinensis seeds by regulating the sucrose and starch pathways with different expression levels.

The Prediction of the Isoelectric Point Value of Peptides and Proteins with a Wide Range of Chemical Modifications

The scale of virtual pKa values for calculating the isoelectric point of peptides and proteins with chemical and post-translational modifications (PTM) is presented. The learning set of pKa values is based on data from 25 experiments of isoelectric focusing of peptides with subsequent mass spectrometric identification (ProteomeXchange accession codes: PXD000065, PXD005410, PXD006291, PXD010006 and PXD017201). In order to enrich the resulting sets with peptides containing modifications the identification of peptides was repeated using raw mass spectrometry data of all datasets. In the final learning set have included peptides satisfying the following conditions: the peptide was found in the fraction with scoring function maximum and maximum peptide abundance; the peptide was found in more than one experiment, and differences of the pI value between experiments was less than 0.15 pH unit. Two variants of the scales were created. In the first variant, pKa values depended only on the residue position relative to the ends of the sequence (N- or C-terminal residue or inside the chain). In the second variant, the effect of neighboring residues was also taken into account. The prediction accuracy of the second variant was higher. The comparison with other methods of pI prediction was carried out. Although the scale was calculated from set containing only peptides, it would be applicable for pI prediction of proteins with and without PTM. The software for prediction of pI values using the resulting pKa scales is available at http://pIPredict3.ibmc.msk.ru.

A method of controllable positive-charged modification of PVDF-CTFE membrane surface based on C–Cl active site

There is a great market demand for positive-charged membrane used to treat wastewater containing positive-charged pollutants. Based on the fact that the C–Cl bond energy in the molecular chain of the PVDF-based poly (vinylidene fluoride-co-chlorotrifluoroethylene) (PVDF-CTFE) copolymer membrane is significantly lower than the C–F bond. Herein, the method of controllable positive-charged modification of PVDF-CTFE membrane surface has been successfully realized by using diethylenetriamine (DETA) a small molecule polyamine compound under mild conditions. The modified membranes were characterized by X-ray photoelectron spectroscopy (XPS), zeta potential and water contact angle, and the best condition for positive-charged modification were explored. The results show that the modified membrane has an excellent positively charged effect, changing from an original membrane which is completely negative to the positive-charged membrane whose isoelectric point value is 7.63, the content of N element is as high as 6.31 at% and the water contact angle decreased by 42.8%. After 7 cycles of treatment of polycationic acrylamide solution, the recovery rate of feed-liquid flux is still above 97.5%, indicating that the ability to resist positively charged pollutants is significantly improved. More importantly, the mechanical strength is not adversely affected. The method proposed in this paper may provide a good idea for the graft modification of chlorinated polymers.

Exploiting the potential of 2D DIGE and 2DE immunoblotting for comparative analysis of crude extract of Trichinella britovi and Trichinella spiralis muscle larvae proteomes

The Trichinella genus poses an interesting puzzle for researchers, having diverged very early in the evolution of the nematodes. The Trichinella spiralis proteome is a cosmopolitan and well-studied model of Trichinella; however, Trichinella britovi also circulates in the sylvatic environment and both species infect humans, resulting in the development of trichinellosis. Few experiments have examined the proteins belonging to the T. britovi proteome. The aim of the present study was to compare the protein expression profiles of crude extracts of T. spiralis and T. britovi muscle larvae using a highly-sensitive two-dimensional differential in-gel electrophoresis (2D DIGE) technique coupled with 2DE immunoblotting. Selected immunoreactive protein spots were then identified by liquid chromatography coupled with mass spectrometry analysis (LC–MS/MS), and their function in Trichinella and the host-parasite interaction was determined by gene ontology analysis. Spots common to both T. spiralis and T. britovi, spots with different expressions between the two and spots specific to each species were labelled with different cyanine dyes. In total, 196 protein spots were found in both proteomes; of these 165 were common, 23 expressed exclusively in T. spiralis and 8 in T. britovi. A comparative analysis of volume ratio values with Melanie software showed that among the common spots, nine demonstrated higher expression in T. spiralis, and 17 in T. britovi. LC–MS/MS analysis of 11 selected spots identified 41 proteins with potential antigenic characteristics: 26 were specific for T. spiralis, six for T. britovi, and eight were found in both proteomes. Gene Ontology analysis showed that the identified T. spiralis proteins possess hydrolytic endopeptidase, endonuclease and transferase activities. Similarly, most of the T. britovi proteins possess catalytic activities, such as lyase, hydrolase, isomerase and peptidase activity. The applied 2D DIGE technique visualized Trichinella spp. protein spots with different molecular weights or isoelectric point values, as well as those with different expression levels. The identified immunoreactive proteins participate in multiple processes associated with host muscle cell invasion and larval adaptation to the host environment. Their reactivity with the host immune system makes them possible candidates for the development of a novel trichinellosis diagnostic test or vaccine against helminthiasis caused by T. spiralis or T. britovi.

Characterization of Cu/Zn-SODs in sympatric species: A comparison of zebra and quagga mussels

Zebra mussels (Dreissena polymorpha) are commonly used as sentinel organisms in freshwater biomonitoring. The displacement of this species by quagga mussels (Dreissena rostriformis bugensis) has raised the issue of their discrimination after field sampling. Even if those two species exhibit specific morphological characteristics, confusion can occur. However, behind the visible phenotypes lie important physiological and metabolic differences that may affect the interpretation of biomarkers in risk assessment. In this study, we present a characterization of Cu/Zn-SODs (superoxide dismutases) in Dreissena spp. with a comparison of activity profiles from individuals originating from North America and Europe. A phylogenetic tree was constructed and dreissenid SOD sequences appear to be closely related to clam species sequences. A specific isoform at an isoelectric point value of 4.95 was observed only for quagga mussels independently of their origin, while zebra mussels exhibited several polymorphic profiles, even for a single sampling site. A comparison of transcript sequences was conducted and the observed differences were highlighted. Alignment of amino acid sequences shows a high level of identity between the two species associated with minor changes in the predicted secondary structures.

The Equilibria in Lipid-Lipoic Acid Systems: Monolayers, Microelectrophoretic and Interfacial Tension Studies.

In this examination, we investigated the effect of lipoic acid (LA) on the properties of biological membrane models (monolayers, bilayers, and liposomes) formed from phosphatidylcholine (PC) or phosphatidylserine (PS) using the Langmuir, microelectrophoresis, and interfacial tension methods. The Langmuir technique allowed us to calculate the π–A isotherms and determine the molecular surface areas of pure and mixed monolayers. Using mathematical equations, we established that LA and the lipids formed complexes at a 1:1 ratio. The interfacial tension method was based on Young and Laplace’s equation. We assumed the formation of a 1:1 complex in the PC–LA system. Using the mathematical relationships, we derived the parameters characterizing the resulting complex, i.e., the surface occupied by the complex and the interfacial tension and stability constant of the formed complex. The microelectrophoretic method was used to determine the dependence of the zeta potential of the lipid membranes as a function of the pH (pH 2 to 10) of the electrolyte solution. The results indicate that modification of PC or PS membranes with LA affects changes in the zeta potential and the isoelectric point values.