Molecular Characterization Of Membrane Proteins Of Schistosoma Mansoni

Schistosomiasis is one of the most prevalent human diseases and is caused by the long-term survival of parasitic blood flukes. Membrane constituents at the surface of these parasites play a major role in parasite metabolism and elaborate sophisticated mechanisms for evasion of the host immune response. Despite this fact, little is known about the molecular properties or functions of individual membrane proteins. This study provides information regarding two polypeptides associated with the apical plasma membrane (APM) of Schistosoma mansoni.;An in vitro system for studying protein phosphorylation in the isolated APM was developed and the protein substrates of endogenous protein kinase activity were described. A 24 kilodalton (kDa) phosphoprotein was characterized in detail. Analysis of the structure of this polypeptide by lectin affinity chromatography, endoglycosidase digestion and phase separation in Triton X-114 demonstrated that the 24 kDa molecule was an integral membrane protein with N-linked oligosaccharides. In addition, the 24 kDa phosphoprotein was shown to be a major APM immunogen by immunoprecipitation with anti-APM antisera and with antibodies affinity purified from the 24 kDa region of preparative Western blots.;In order to isolate cDNA clones encoding APM polypeptides, comprehensive cDNA expression libraries were constructed in {dollar}\lambda{dollar} bacteriophage vectors and were screened with anti-APM antisera. Two cDNA clones were isolated and their nucleotide sequences determined. One cDNA was 141 base pairs in length and was shown to encode antigenic determinants shared with the 24 kDa phosphoprotein antigen. This cDNA was not full length since the homologous mRNA was approximately 800 residues in length. A possible open reading frame from this cDNA however, contained a signal for N-linked glycosylation. A second near full-length cDNA of 2621 base pairs was also cloned and sequenced. An open reading frame deduced from this cDNA predicted a protein of 702 amino acids with a molecular weight of 76 kDa. The deduced protein sequence was shown to be similar to the known sequences of vertebrate calpains.;This study provides the basis for a detailed structure/function analysis of the membrane-associated polypeptides encoded by the cloned cDNAs

Spatial distribution of selected chemical cell wall components in the embryogenic callus of Brachypodium distachyon

Brachypodium distachyon L. Beauv. (Brachypodium) is a species that has become an excellent model system for gaining a better understanding of various areas of grass biology and improving plant breeding. Although there are some studies of an in vitro Brachypodium culture including somatic embryogenesis, detailed knowledge of the composition of the main cell wall components in the embryogenic callus in this species is missing. Therefore, using the immunocytochemical approach, we targeted 17 different antigens of which five were against the arabinogalactan proteins (AGP), three were against extensins, six recognised pectic epitopes and two recognised hemicelluloses. These studies were complemented by histological and scanning electron microscopy (SEM) analyses. We revealed that the characteristic cell wall components of Brachypodium embryogenic calli are AGP epitopes that are recognised by the JIM16 and LM2 antibodies, an extensin epitope that is recognised by the JIM11 antibody and a pectic epitopes that is recognised by the LM6 antibody. Furthermore, we demonstrated that AGPs and pectins are the components of the extracellular matrix network in Brachypodium embryogenic culture. Additionally, SEM analysis demonstrated the presence of an extracellular matrix on the surface of the calli cells. In conclusion, the chemical compositions of the cell walls and ECMSN of Brachypodium callus show spatial differences that correlate with the embryogenic character of the cells. Thus, the distribution of pectins, AGPs and hemicelluloses can be used as molecular markers of embryogenic cells. The presented data extends the knowledge about the chemical composition of the embryogenic callus cells of Brachypodiu

Effect of Pseudomonas moorei KB4 Cells’ Immobilisation on Their Degradation Potential and Tolerance towards Paracetamol

Pseudomonas moorei KB4 is capable of degrading paracetamol, but high concentrations of this drug may cause an accumulation of toxic metabolites. It is known that immobilisation can have a protective effect on bacterial cells; therefore, the toxicity and degradation rate of paracetamol by the immobilised strain KB4 were assessed. Strain KB4 was immobilised on a plant sponge. A toxicity assessment was performed by measuring the concentration of ATP using the colony-forming unit (CFU) method. The kinetic parameters of paracetamol degradation were estimated using the Hill equation. Toxicity analysis showed a protective effect of the carrier at low concentrations of paracetamol. Moreover, a pronounced phenomenon of hormesis was observed in the immobilised systems. The obtained kinetic parameters and the course of the kinetic curves clearly indicate a decrease in the degradation activity of cells after their immobilisation. There was a delay in degradation in the systems with free cells without glucose and immobilised cells with glucose. However, it was demonstrated that the immobilised systems can degrade at least ten succeeding cycles of 20 mg/L paracetamol degradation. The obtained results indicate that the immobilised strain may become a useful tool in the process of paracetamol degradation

Long Term Friction: from Stick-Slip to Stable Sliding

We have devised an original laboratory experiment where we investigate the frictional behaviour of a single crystal salt slider over a large number of deformation cycles. Because of its physical properties, salt, a surrogate for natural faults, allows for friction and plastic deformation and pressure solution creep to be efficient on the same timescale. During the same experiment, we observe a continuous change of the frictional behaviour of the slider under constant conditions of stiffness, temperature and loading velocity. The stick-slip regime is progressively vanishing, eventually reaching the stable sliding regime. Concomitantly, the contact interface, observed under the microscope, develops a striated morphology with contact asperities increase in length and width, arguing for an increase in the critical slip distance dc. Complementary experiments including velocity jumps show that the frictional parameters of the rate and state friction law, a and b, progressively vanish with the cumulative slip. In our experimental conditions, the ultimate stage of friction is therefore rate and state independent.Comment: 10 pages; 4 figures; 1 Tabl

Supplementary data for article: Solarz, D., Witko, T., Karcz, R., Malagurski, I., Ponjavić, M., Levic, S., Nešić, A., Guzik, M., Savić, S.,& Nikodinović-Runić, J.. (2023). Biological and physiochemical studies of electrospun polylactid/polyhydroxyoctanoate PLA/ P(3HO) scaffolds for tissue engineering applications. in RSC Advances, 13(34), 24112-24128. https://doi.org/10.1039/D3RA03021K

Polyhydroxyoctanoate, as a biocompatible and biodegradable biopolymer, represents an ideal candidate for biomedical applications. However, physical properties make it unsuitable for electrospinning, currently the most widely used technique for fabrication of fibrous scaffolds. To overcome this, it was blended with polylactic acid and polymer blend fibrous biomaterials were produced by electrospinning. The obtained PLA/PHO fibers were cylindrical, smaller in size, more hydrophilic and had a higher degree of biopolymer crystallinity and more favorable mechanical properties in comparison to the pure PLA sample. Cytotoxicity evaluation with human lung fibroblasts (MRC5 cells) combined with confocal microscopy were used to visualize mouse embryonic fibroblasts (MEF 3T3 cell line) migration and distribution showed that PLA/PHO samples support exceptional cell adhesion and viability, indicating excellent biocompatibility. The obtained results suggest that PLA/PHO fibrous biomaterials can be potentially used as biocompatible, biomimetic scaffolds for tissue engineering applications.Supplementary material for:[https://doi.org/10.1039/D3RA03021K]Related to published version: [https://imagine.imgge.bg.ac.rs/handle/123456789/2059

Biological and physiochemical studies of electrospun polylactid/polyhydroxyoctanoate PLA/ P(3HO) scaffolds for tissue engineering applications

Polyhydroxyoctanoate, as a biocompatible and biodegradable biopolymer, represents an ideal candidate for biomedical applications. However, physical properties make it unsuitable for electrospinning, currently the most widely used technique for fabrication of fibrous scaffolds. To overcome this, it was blended with polylactic acid and polymer blend fibrous biomaterials were produced by electrospinning. The obtained PLA/PHO fibers were cylindrical, smaller in size, more hydrophilic and had a higher degree of biopolymer crystallinity and more favorable mechanical properties in comparison to the pure PLA sample. Cytotoxicity evaluation with human lung fibroblasts (MRC5 cells) combined with confocal microscopy were used to visualize mouse embryonic fibroblasts (MEF 3T3 cell line) migration and distribution showed that PLA/PHO samples support exceptional cell adhesion and viability, indicating excellent biocompatibility. The obtained results suggest that PLA/PHO fibrous biomaterials can be potentially used as biocompatible, biomimetic scaffolds for tissue engineering applications.Related to supplementary material: [https://imagine.imgge.bg.ac.rs/handle/123456789/2060

Gene encoding a deubiquitinating enzyme is mutated in artesunate- and chloroquine-resistant rodent malaria parasites§

Artemisinin- and artesunate-resistant Plasmodium chabaudi mutants, AS-ART and AS-ATN, were previously selected from chloroquine-resistant clones AS-30CQ and AS-15CQ respectively. Now, a genetic cross between AS-ART and the artemisinin-sensitive clone AJ has been analysed by Linkage Group Selection. A genetic linkage group on chromosome 2 was selected under artemisinin treatment. Within this locus, we identified two different mutations in a gene encoding a deubiquitinating enzyme. A distinct mutation occurred in each of the clones AS-30CQ and AS-ATN, relative to their respective progenitors in the AS lineage. The mutations occurred independently in different clones under drug selection with chloroquine (high concentration) or artesunate. Each mutation maps to a critical residue in a homologous human deubiquitinating protein structure. Although one mutation could theoretically account for the resistance of AS-ATN to artemisinin derivates, the other cannot account solely for the resistance of AS-ART, relative to the responses of its sensitive progenitor AS-30CQ. Two lines of Plasmodium falciparum with decreased susceptibility to artemisinin were also selected. Their drug-response phenotype was not genetically stable. No mutations in the UBP-1 gene encoding the P. falciparum orthologue of the deubiquitinating enzyme were observed. The possible significance of these mutations in parasite responses to chloroquine or artemisinin is discussed