Secondary substrate binding strongly affects activity and binding affinity of Bacillus subtilis and Aspergillus niger GH11 xylanases

The secondary substrate binding site (SBS) of Bacillus subtilis and Aspergillus niger glycoside hydrolase family 11 xylanases was studied by site-directed mutagenesis and evaluation of activity and binding properties of mutant enzymes on different substrates. Modification of the SBS resulted in an up to three-fold decrease in the relative activity of the enzymes on polymeric versus oligomeric substrates and highlighted the importance of several amino acids in the SBS forming hydrogen bonds or hydrophobic stacking interactions with substrates. Weakening of the SBS increased K(d) values by up to 70-fold in binding affinity tests using natural substrates. The impact that modifications in the SBS have both on activity and on binding affinity towards polymeric substrates clearly shows that such structural elements can increase the efficiency of these single domain enzymes on their natural substrates.     

Effects of phenolic monomers on growth of Acidothermus cellulolyticus

Previous studies on biological pretreatment of switchgrass by solid-state fermentation with Acidothermus cellulolyticus 11B have shown that inhibitory compounds prevent growth on untreated switchgrass. A. cellulolyticus was grown in liquid medium containing cellobiose with phenolic monomers added to determine if the phenolic compounds are one possible source of inhibition. Cinnamic acid derivatives (trans-p-coumaric, trans-ferulic, and hydrocinnamic acids), hydroxybenzoic acids (p-hydroxybenzoic, syringic, and vanillic acids), benzaldehydes (vanillin and p-hydroxybenzaldehyde), and condensed tannin monomers (catechin and epicatechin) were tested at levels up to 20 mM. All compounds exhibited a dose-response relationship and strongly inhibited growth at 20 mM. trans-p-Coumaric acid was found to be the strongest inhibitor of A. cellulolyticus growth, with a specific growth rate of 0.004 h(-1) at 1 mM (0.18 h(-1) without phenolic monomer). GC-MS and HPLC methods were used to confirm the presence of these phenolic compounds in switchgrass and measure the amounts extracted using different conditions. The amounts of phenolic compounds measured were found to be higher than the threshold for growth inhibition. Leaching with water at 55°C was inefficient at removing bound phenolics, whereas NaOH treatment improved efficiency. Phenolic compounds spiked into alkaline pretreated switchgrass were also found to inhibit growth of A. cellulolyticus in solid-state fermentation. However, addition of insoluble polyvinylpolypyrrolidone (PVPP) to switchgrass improved growth of A. cellulolyticus in liquid cultures, providing a possible approach for alleviating microbial inhibition due to phenolic compounds in lignocellulose.     

Sublethal effects of essential oil of Cinnamomum zeylanicum Blume on life expectancy (ex) and age-specific fertility (mx) of two-spotted spider mite,Tetranychus urticae Koch (Acari: Tetranychidae)

Two-spotted spider mite, Tetranychus urticae Koch, is one of the most important pests of agricultural products that have a global distribution. Now, the control is dependent on the use of chemical pesticides. The effects of the sublethal concentrations (LC₁₀ and LC₂₅) of the essential oil of Cinnamomum zeylanicum were evaluated on some parameters of the life table of the pest, at the constant temperature of 30?°C, relative humidity of 40?±?5% and photoperiod of 16L: 8D. Our results showed that the essential oil of cinnamon is effective on female adult stage. Lethal concentration at which 50% mortality (LC₅₀) for the essential oils is from C. zeylanicum on female adult mite was 23.39?μl/L air. Sublethal concentrations of the essential oil of C. zeylanicum also impair the natural biology of the mite. Concentrations of sublethal of essential oil decreased age-specific fertility (m_x) of T. urticae compared with the control. Sublethal concentrations (LC₁₀ and LC₂₅) have reduced the life expectancy (e_x) in egg stage. Sublethal effects of the essential oil of C. zeylanicum and its impact on T. urticae management are discussed.     

Mesenchymal stem cells alter the frequency and cytokine profile of natural killer cells in abortion-prone mice

Natural killer cells, which play a pivotal role in the establishment and maintenance of normal pregnancy, are the most abundant leukocytes at the fetomaternal interface that their subsets frequencies and cytokine profile are influential factors in the preservation of the decidual tolerogenic microenvironment. Any imbalance in NK cells' frequency and functions could be associated with pregnancy failure. Mesenchymal stem cells (MSCs) are shown to have immunomodulatory effects on NK cells and their cytokine profile. The purpose of this study is to evaluate the impact of MSCs therapy on the cytokine profiles and subpopulations of NK cells in a murine model of recurrent pregnancy loss. Adipose-derived MSCs were injected intraperitoneally to the abortion-prone mice on Day 4.5 of gestation. The abortion rate was determined after MSCs administration and the frequency and cytokine profiles of the different subsets of NK cells were determined using the flow cytometry. Our results showed that, in abortion-prone mice, the frequency of CD49b⁺ NK cells was significantly higher than normal pregnant mice that decreased after therapy. We also demonstrated that MSCs downregulated the production of IFN-γ and upregulated IL-4 and IL-10 production by uNK cells. These findings indicate that MSCs can decrease the infiltration of CD49b⁺ NK cells to the fetomaternal interface and modulate the cytokine profile of NK cells from inflammatory to tolerogenic profile and thereby improve the tolerogenic microenvironment at the fetomaternal interface in benefit of pregnancy maintenance.     

Lymphopenia in COVID‐19: Therapeutic opportunities

Severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) is uncontrollably spread all over the world. The host immune responses strongly try to confront it with all the potential cells and cytokines. With chronically condition of SARS‐CoV‐2, natural killer cells and T cells become exhausted and decreasing their count leads to lymphopenia. Inability to eradicate the infected organ makes hyperinitiation of the immune system, which releases the excessive inflammatory cytokines to compensate the exhausted one as well as the low lymphocytes counts; it consequently leads to the cytokine storm syndrome. These mechanisms and the potential therapeutic targeting are discussed in this paper.     

Polarization Controlling of Multi Resonant Graphene-Based Microstrip Antenna

Plasmonics - In this paper, a graphene-based patch antenna is proposed. The antenna structure is designed so that each of the various antenna sections affected by chemical potential changes can...     

Enhancement of extracellular bispecific anti‐MUC1 nanobody expression in E. coli BL21 (DE3) by optimization of temperature and carbon sources through an autoinduction condition

Escherichia coli is one of the most suitable hosts for production of antibodies and antibody fragments. Antibody fragment secretion to the culture medium improves product purity in cell culture and diminishes downstream costs. In this study, E. coli strain BL21 (DE3) harboring gene encoding bispecific anti‐MUC1 nanobody was selected, and the autoinduction methodology for expression of bispecific anti‐MUC1 nanobody was investigated. Due to the replacement of IPTG by lactose as inducer, less impurity and toxicity in the final product were observed. To increase both intracellular and extracellular nanobody production, initially, the experiments were performed for the key factors including temperature and duration of protein expression. The highest amount of nanobody was produced after 21 h at 33°C. The effect of different carbon sources, glycerol, glucose, lactose, and glycine as a medium additive at optimum temperature and time were also assessed by using response surface methodology. The optimized concentrations of carbon sources were obtained as 0.75% (w/v), 0.03% (w/v), 0.1% (w/v), and 0.75% (w/v) for glycerol, glucose, lactose, and glycine, respectively. Finally, the production of nanobody in 2 L fermenter under the optimized autoinduction conditions was evaluated. The results show that the total titer of 87.66 µg/mL anti‐MUC1 nanobody, which is approximately seven times more than the total titer of nanobody produced in LB culture medium, is 12.23 µg/L .     

Contribution of DNA methylation and EZH2 in SRBC down-regulation in gastric cancer

Molecular Biology Reports - Gastric cancer (GC), a high mortality malignancy, is induced by genetic and epigenetic factors. DNA and histone methylation play critical roles in tumor suppressor genes...     

Deoxycholic acid causes DNA damage while inducing apoptotic resistance through NF-κB activation in benign Barrett's epithelial cells

Gastroesophageal reflux is associated with adenocarcinoma in Barrett's esophagus, but the incidence of this tumor is rising, despite widespread use of acid-suppressing medications. This suggests that refluxed material other than acid might contribute to carcinogenesis. We looked for potentially carcinogenetic effects of two bile acids, deoxycholic acid (DCA) and ursodeoxycholic acid (UDCA), on Barrett's epithelial cells in vitro and in vivo. We exposed Barrett's (BAR-T) cells to DCA or UDCA and studied the generation of reactive oxygen/nitrogen species (ROS/RNS); expression of phosphorylated H2AX (a marker of DNA damage), phosphorylated IkBα, and phosphorylated p65 (activated NF-κB pathway proteins); and apoptosis. During endoscopy in patients, we took biopsy specimens of Barrett's mucosa before and after esophageal perfusion with DCA or UDCA and assessed DNA damage and NF-κB activation. Exposure to DCA, but not UDCA, resulted in ROS/RNS production, DNA damage, and NF-κB activation but did not increase the rate of apoptosis in BAR-T cells. Pretreatment with N-acetyl-l-cysteine (a ROS scavenger) prevented DNA damage after DCA exposure, and DCA did induce apoptosis in cells treated with NF-κB inhibitors (BAY 11-7085 or AdIκB superrepressor). DNA damage and NF-κB activation were detected in biopsy specimens of Barrett's mucosa taken after esophageal perfusion with DCA, but not UDCA. These data show that, in Barrett's epithelial cells, DCA induces ROS/RNS production, which causes genotoxic injury, and simultaneously induces activation of the NF-κB pathway, which enables cells with DNA damage to resist apoptosis. We have demonstrated molecular mechanisms whereby bile reflux might contribute to carcinogenesis in Barrett's esophagus.