Biotechnology and applied biochemistry最新文献

The effects of 180, 210, and 230°C reaction temperatures on the structural and magnetic properties of synthesized iron sulfide nanoparticles were studied. The Rietveld refinement analysis result of the X-ray diffraction data indicated that greigite was the dominant phase in all samples. The sample was prepared at 210°C for 18 h and had a greater wt% ratio of the greigite phase. The crystallite and particle sizes increased with increasing reaction temperatures. Scanning electron microscope images confirmed the presence of aggregation of synthesized rod-shaped nanoparticles. The magnetic hysteresis curves of all samples showed ferromagnetic behavior at room temperature. The magnetic saturation of three samples increases with increased reaction temperature, but the coercive force has the opposite behavior. Antioxidant activity and cytotoxicity of the sample synthesized at 210°C were investigated. This sample killed cancer cells at relatively moderate and high concentrations with high viability of normal cells, demonstrating the sample's suitability for use in killing cancer cells while avoiding normal cells.

Maximizing the recombinant protein yield necessitates optimizing the production medium. This can be done using a variety of methods, including the conventional “one-factor-at-a-time” approach and more recent statistical and mathematical methods such as artificial neural network (ANN), genetic algorithm, etc. Every approach has advantages and disadvantages of its own, yet even when a technique has flaws, it is nevertheless used to get the best results. Here, one categorical variable and four numerical parameters, including post-induction time, inducer concentration, post-induction temperature, and pre-induction cell density, were optimized using the 232 experimental assays of the central composite design. The direct and indirect effects of factors on the yield of anti-epithelial cell adhesion molecule extracellular domain fragment antibody were examined using statistical methods. The analysis of variance results indicate that the response surface methodology (RSM) model is effective in predicting the amount of produced single-chain fragment variable (p-value = 0.0001 and R² = 0.905). For ANN modeling, the evaluation using normalized root mean square error (NRMSE) and R² values shows a good fit (R² = 0.942) and accurate predictions (NRMSE = 0.145). The analysis of error parameters and R² of a dataset, which contained 30 data points randomly selected from the complete dataset, showed that the ANN model had a higher R² value (0.968) compared to the RSM model (0.932). Furthermore, the ANN model demonstrated stronger predictive ability with a lower NRMSE (0.048 vs. 0.064). Induction at the cell density of 0.7 and an isopropyl β-D-1-thiogalactopyranoside concentration of 0.6 mM for 32 h at 30°C in BW25113 was the ideal culture condition leading to the protein yield of 259.51 mg/L. Under the optimum conditions, the output values predicted by the ANN model (259.83 mg/L) were more in line with the experimental data (259.51 mg/L) than the RSM (276.13 mg/L) expected value. This outcome demonstrated that the ANN model outperforms the RSM in terms of prediction accuracy.

Rapid control of the content of Parkinson's drugs in biological fluids and pharmaceutical formulations is of great importance because changes in the concentration of these drugs affect their bioavailability and biopharmaceutical properties. Therefore, we presented a simple and convenient method for the ratiometric detection of carbidopa and levodopa for carbon dots (CDs) dual-fluorescent emission. Dual-emission CDs were prepared from chitosan using a microwave method, following which the surface was chemically modified with terephthalaldehyde. CDs had two strong well-separated peaks at 445 and 510 nm. The relative measurement of carbidopa and levodopa was based on the static extinction of CDs at 445 nm and increase at 510 nm, respectively. The linear range for carbidopa measurement was 2.5–300 nM, with a limit of detection (LOD) of 2.1 nM, and a relative standard deviation (RSD) of 1.68%. Further, the linear range for levodopa measurement was equal to 3.0–400 nM, with LOD and RSD% of 2.8 nM and 3.5%, respectively. Also, selectivity of ratiometric sensor in the presence of interferences was investigated, which showed that the recovery of carbidopa and levodopa in serum and urine samples has changed between 96.80% and 116.24% with RSD% 0.11–0.77. CDs also provided good results for the determination of carbidopa and levodopa in real samples, and had high selectivity in the presence of possible interferences.

Outer surface/membrane and virulent secretory proteins are primarily crucial for pathogenesis. Secreted and outer membrane hydrolases of many pathogens play an important role in attenuating the host immune system. Leptospira expresses many such proteins, and few have been characterized to display various roles, including host immune evasion. However, identification, classification, characterization, and elucidation of the possible role of Leptospira’s outer membrane and secretory hydrolases have yet to be explored. In the present study, we used bioinformatics tools to predict exported proteins from the pathogenic Leptospira proteome. Moreover, we focused on secretory and outer membrane putative hydrolases from the exported proteins to generate a deeper understanding. Our analysis yielded four putative outer/secretory hydrolases, LIC_10995, LIC_11183, LIC_11463, and LIC_12988, containing α/β hydrolase fold and displayed similarity with lipase motif. Moreover, their conservation analysis of the predicted hydrolases across the spectrum of different Leptospira species showed high clustering with the pathogenic species. Outer membrane and secretory proteins with lipolytic activity may have a role in pathogenesis. This is the first bioinformatics analysis of secretory and outer membrane α/β hydrolases from leptospiral species. However, experimental studies are indeed required to unravel this possibility.

Although a series of studies confirm the bioactivities of hederagenin and its glycosides, their synergistic effects and potential mechanisms are still worthy of further exploration. This work investigated the synergistic cytotoxicity and in vitro antioxidant activity of hederagenin and hederagenin 28-O-β-d-glucopyranoside (28-Glc-hederagenin). Hederagenin and 28-Glc-hederagenin inhibited HeLa cell growth and their combination further strengthened this effect. The combination of hederagenin and 28-Glc-hederagenin significantly increased the rate of apoptotic cells, suggesting the presence of a synergistic effect between the two substances. This combination also enhanced in vitro antioxidant activity compared with individual treatments. A network pharmacology and molecular docking-based approach was performed to explore the underlying mechanisms of hederagenin and 28-Glc-hederagenin against cervical cancer and oxidant damage. This work identified 18 related Kyoto Encyclopedia of Genes and Genome pathways, 202 related biological process terms, 17 related CC terms, and 35 related molecular function terms and then revealed 30 nodes and 196 edges. Subsequently, two highly connected clusters and the top four targets were identified. Molecular docking showed potent binding affinity of hederagenin and 28-Glc-hederagenin toward core targets associated with both cervical cancer and oxidant damage. This work may provide scientific basis for the combined use of hederagenin and its glycosides as dietary supplements.

The identification of novel acetylcholinesterase inhibitors holds significant relevance in the treatment of Alzheimer's disease (AD), the prevailing form of dementia. The exploration of alternative inhibitors to the conventional acetylcholinesterase inhibitors is steadily gaining prominence. Quinones, categorized as plant metabolites, represent a specific class of compounds. In this study, the inhibitory effects of various naphthoquinone derivatives, along with anthraquinone and its derivatives, on the acetylcholinesterase (AChE) enzyme were investigated for this purpose. An in vitro investigation was conducted to examine the effects of these compounds in order to clarify the possible mechanism of inhibition in the interaction between the enzyme and chemicals. In addition, an in silico investigation was carried out to understand the conceivable inhibitor binding process to the enzyme's active site. The acquired outcomes corroborated the in vitro results. The AChE enzyme was found to be effectively inhibited by both naphthoquinones and anthraquinones, with inhibition constant (K_I) values ranging from 0.014 to 0.123 μM (micormolar). The AChE enzyme was inhibited differently by this quinone and its derivatives. Although derivatives of naphthoquinone and anthraquinone exhibited a competitive inhibitory effect, derivatives of anthraquinone exhibited a noncompetitive inhibition effect. Furthermore, because it had the lowest K_I value of any of these substances, 1,5-dihydroxyanthraquinone (1c) was shown to be the most potent inhibitor. The findings will add to the body of knowledge on the creation of fresh, potent, and successful treatment approaches.

Purpose of this article is to study the possible direct antiviral effect of “Armenikum” on SARS-CoV-2, conduct an in vitro study on the SARS-CoV-2 encephalomocarditis virus, and an in vivo study on the Syrian hamster model. Human coronavirus SARS-CoV-2 (delta strain) was used as the virus. Two groups of four-specimen hamsters were used to study the therapeutic activity of the drug during 48 h after infecting. One group of hamsters served as positive control and was infected with the virus at a similar dose as experimental one and was used as a control of pathology induced by the viral infection till the end of the experiment. Another group of hamsters (four of them) was injected physiological solution and was used as a control. The Syrian hamsters underwent a clinical blood test and computed tomography. “Armenikum” in the form of an injection has a significant antiviral effect on the human coronavirus SARS-CoV-2, credibly reducing the titers of the virus and the time of its elimination from the Syrian hamsters, significantly mitigating the viral infection. “Armenikum” in the form of an injection drug almost completely removes the pathological effect of the virus in the lungs of the hamsters.

Glutamate dehydrogenase (GDH) participates in the energy metabolism of proteins and the synthesis of metabolites important for the organism. In this study, GDH enzyme was purified from the liver of rainbow trout (Oncorhynchus mykiss) by 2',5'-ADP Sepharose 4B affinity chromatography in one step. As a result of this purification process, GDH enzyme was purified 171-fold with 5.83 U/mg protein-specific activity. The characterization experiments presented that the storage stability of the purified GDH enzyme was determined as −80°C; optimum temperature 40°C; it was determined that the optimum ionic strength was 100 mM phosphate buffer and the optimum pH was 8.00. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and PAGE studies showed that the natural molar mass of the purified GDH enzyme was 346.74 kDa, and the molar mass of its subunits was 53.71 kDa. K_m and V_max values for substrates and coenzymes of GDH enzyme purified from rainbow trout liver were calculated, and the lowest K_m value was found in NAD⁺ (1.86 mM) and the highest V_max value in NH₄⁺ (1.79 U/mL). The effects of some metal ions, vitamins, and solvents on the activity of the purified GDH enzyme were investigated and also IC₅₀ values and inhibition types. The metal ion with the lowest IC₅₀ value is Ag⁺ (8.65 ± 1.68 μM), and the vitamin is B₆ (0.77 ± 0.04 mM). The binding affinities of inhibitors were investigated with molecular docking, based on the conformational state of GDH.

Breast cancer (BC) is the most common cancer type and the fifth leading cause of cancer-related deaths. The primary goals of BC treatment are to remove the tumor and prevent metastasis. Despite advances in BC treatment, more effective therapies are required. miRNAs can regulate many targets involved in biological processes and tumor progression; these molecules have emerged as a promising cancer treatment strategy. In the present study, we investigated the effects of miR-99a and miR-143 in single expression plasmids for BC inhibition. In this study, the precursor structure of miRNAs in the expression vector pEGFP-N1 entered single and double states, and MCF7 and T47D cells were transfected. The miRNAs expression level after transfection was then measured using qPCR. The MultiMiR package was used to obtain predicted and validated miRNA targets. MTT assay, qRT-PCR, migration test, and flow cytometry were used to assess the effect of miRNA and gene modulation. The qPCR results revealed that miRNA constructs were significantly expressed after the transfection of both cell lines. The biological function of miRNAs showed that upregulation of miR-99a and miR-143 in any of the two selected BC cells inhibited their proliferation and migration rate, significantly inducing apoptosis (p < 0.01). Also, miR-99a/miR-143 co-treatment has a synergistic anticancer effect in cancer cells via Akt1 and CDK6 targeting. These findings suggest that miR-99a/miR-143 plays synergistic regulatory roles in BC, possibly via a shared signaling pathway, providing a therapeutic strategy for BC treatment.

Recently, concern has been raised about the spread of human mpox virus, and the demand for rapid detection reagents for mpox virus has increased. This study aims to establish a time-resolved fluorescence immunochromatography (TRFICO) method for qualitative/quantitative detection of mpox virus. A double-antibody sandwich TRFICO method was optimized and established using mpox recombinant fusion antigen and its paired monoclonal antibody. The test performance of the method was evaluated using mpox fusion antigen and control serum, including sensitivity, linearity range, specificity, precision, and reference interval. We successfully established a TRFICO method for qualitative/quantitative detection of mpox antigen, its linearity range 0–100 ng/mL, analytical sensitivity 0.017 ng/mL, and reference intervals greater than 0.045 ng/mL. No cross-reaction was detected with various poxvirus and clinical negative controls, with good specificity. All average recoveries of the intra- and inter-batch ranged from 81.33% to 97.83%, and all CVs were below 10%. Additionally, the TRFICO strips can be stably stored at 37°C for 7 days without significant changes in the fluorescence intensity. This TRFICO method, with high sensitivity, linearity range, specificity, precision, and stability with 16-min detection time, provides a new option for qualitative/quantitative and convenient testing of mpox virus.