Biotechnology Reports最新文献

Bacillus thuringiensis parasporin-2 (PS2Aa1 or Mpp46Aa1) selectively destroys human cancer cells, making it a promising anticancer agent. PS2Aa1 protoxin expression in Escherichia coli typically results in inclusion bodies that must be solubilized and digested by proteinase K to become active. Here, maltose-binding protein (MBP) was fused to the N-terminus of PS2Aa1, either full-length (MBP-fPS2) or truncated (MBP-tPS2), to increase soluble protein expression in E. coli and avoid solubilization and proteolytic activation. Soluble MBP-fPS2 and MBD-tPS2 proteins were produced in E. coli and purified with endotoxin levels below 1 EU/μg. MBP-fPS2 was cytotoxic against T cell leukemia MOLT-4 and Jurkat cell lines after proteinase-K digestion. However, MBP-tPS2 was cytotoxic immediately without MBP tag removal or activation. MBP-tPS2′s thermal stability also makes it appropriate for bioproduction and therapeutic applications.

Deep eutectic solvents (DES) are green alternatives for conventional solvents. They have gained attention for their potential to extract valuable compounds from biomass, such as seaweed. In this framework, a case study was developed to assess the feasibility of pressure-driven membrane processes as an efficient tool for the recovery of deep eutectic solvents and targeted biomolecules. For this purpose, a mixture composed of the DES choline chloride – ethylene glycol (ChCl-EG) 1:2, water and alginate was made to mimic a DES extraction from seaweed. An integrated separation process design was proposed where ultrafiltration-diafiltration-nanofiltration (UF-DF-NF) was coupled. UF and DF were found to be effective for the separation of alginate with an 85 % yield. DES was likewise recovered by 93 %, proving the membrane filtrations’ technical feasibility. The NF performance to separate the DES from the water, for its recycling, laid by a 45 %-50 % retention and a final concentrated DES solution of 18 %(v/v).

Plant-based manufacturing has the advantage of post-translational modifications. While plant-specific N-glycans have been associated with allergic reactions, their effect on the specific immune response upon vaccination is not yet understood. In this study, we produced an RBD-Fc subunit vaccine in both wildtype (WT) and glycoengineered (∆XF) Nicotiana benthamiana plants. The N-glycan analysis: RBD-Fc carrying the ER retention peptide mainly displayed high mannose. When produced in WT RBD-Fc displayed complex-type (GnGnXF) N-glycans. In contrast, ∆XF plants produced RBD-Fc with humanized complex N-glycans that lack potentially immunogenic xylose and core fucose residues (GnGn). The three recombinant RBD-Fc glycovariants were tested. Immunization with any of the RBD-Fc proteins resulted in a similar titer of anti-RBD antibodies in mice. Likewise, antisera from subunit RBD-Fc vaccines also demonstrated comparable neutralization against SARS-CoV-2. Thus, we conclude that N-glycan modifications of the RBD-Fc protein have no impact on their capacity to activate immune responses and induce neutralizing antibody production.

Background

Hepatocellular carcinoma (HCC) is one of the most serious types of cancer that accounts for numerous cancer deaths worldwide. HCC is poorly prognosed and is a highly chemotherapy-resistant tumor. Therefore, new treatments are urgently needed. Exopolysaccharides (EPS-1) produced from the novel Bacillus sonorensis strain was found to exhibit chemopreventive effects against cancer.

Objective

Evaluating the anti-cancer cytotoxic effect of exopolysaccharides (EPS-1) produced by the newly studied Bacillus sonorensis strain SAmt2.

Methods

The cytotoxic activity was investigated through cell cycle, apoptosis, and autophagy analyses using flow cytometry technique. Also, the effect of EPS-1 on Huh7 release of COX-2 was examined using ELISA.

Results

Our results revealed that EPS-1exhibit an anti-proliferative effect on Huh7 cells through decreasing the percentage of cells at the S-phase and G2 phase, while increasing the cell population at the sub-G1 and G1 phases. Apoptosis analysis showed that EPS-1 increased necrotic and apoptotic cell fractions in EPS-1 treated Huh7. In addition, it induced significant autophagic cell death in the Huh7.Finally, antiproliferative and apoptosis induction results were supportedby ELISA assay results where the protein level of COX-2 was declined.

Conclusion

: In conclusion, EPS-1 derived from B. sonorensis SAmt2, is a promising proliferation inhibitor of Huh7 cells with potential anticancer effects.

The present study investigates S. cumini seed extracts which are considered as a promising and valuable source of bioactive compounds were prepared using different solvents such as methanol, ethanol, petroleum ether, acetone, chloroform, and diethyl ether. Among these solvents, methanol exhibited the highest extraction with a yield of 42 %. HPLC analysis revealed the highest concentration of quercetin flavonoids (49.62 mg/gm) in the methanolic S. cumini seed extract. Thus, the current work deals with the MgONPs synthesis through a biological approach using different S. cumini seed extracts. In vitro anti-oxidant properties were evaluated, which showed an IC₅₀ value of 22.46 μg/mL for MgONPs synthesized from methanolic extract, surpassing the anti-oxidant potency of ascorbic acid by threefold. By leveraging the rich repository of bioactive compounds found within S. cumini seed extract, this study presents a novel approach to MgONPs synthesis. Exploring the symbiotic relationship between S. cumini seed extract and MgONPs, this research elucidates the pivotal role of bioactive compounds in guiding the formation and properties of nanostructures. Further anti-microbial studies on MgONPs from methanolic S. cumini seed extract were conducted against four different bacterial strains (Escherichia coli, Bacillus subtilis, Staphylococcus aureus, and S. typhimurium), revealing potent anti-microbial activity with 5.3 mm of inhibition for 100 µl against S. typhimurium. These findings suggest that S. cumini is a source of bioactive compounds responsible for the successful synthesis of MgONPs. Characterization studies of MgONPs were also carried out using UV–vis spectroscopy, FTIR, SEM, XRD, DSC and HPLC.

With the progress in two distinct areas of nanotechnology and aptamer identification technologies, the two fields have merged to what is known as aptamer nanotechnology. Aptamers have varying properties in the biomedical field include their small size, non-toxicity, ease of manufacturing, negligible immunogenicity, ability to identify a wide range of targets, and high immobilizing capacity. Nevertheless, aptamers can utilize the distinct characteristics offered by nanomaterials like optical, magnetic, thermal, electronic properties to become more versatile and function as a novel device in diagnostics and therapeutics. This engineered aptamer conjugated nanomaterials, in turn provides a potentially new and unique properties apart from the pre-existing characteristics of aptamer and nanomaterials, where they act to offer wide array of applications in the biomedical field ranging from drug targeting, delivery of drugs, biosensing, bioimaging. This review gives comprehensive insight of the different aptamer conjugated nanomaterials and their utilization in biomedical field. Firstly, it introduces on the aptamer selection methods and roles of nanomaterials offered. Further, different conjugation strategies are explored in addition, the class of aptamer conjugated nanodevices being discussed. Typical biomedical examples and studies specifically, related to drug delivery, biosensing, bioimaging have been presented.

Dengue virus (DENV), transmitted by mosquitoes, is classified into four serotypes (DENV1-4) and typically causes mild, self-limiting symptoms upon initial infection. However, secondary infection can lead to severe symptoms due to antibody-dependent enhancement (ADE). To address this, anti-DENV antibodies are being developed with the goal of neutralizing infection without ADE activity. Previous attempts using a 54_hG1 antibody from CHO-K1 mammalian cells resulted in ADE induction, increasing viral infection. This study aimed to express the D54 monoclonal antibody in Nicotiana benthamiana. The plant-produced antibody had a similar neutralizing profile to the previous 54_hG1 antibody. Notably, the ADE activities of the plant-derived antibody were successfully eliminated, with no sign of viral induction. These findings suggest that N. benthamiana could be a source of therapeutic DENV antibodies. The method offers several advantages, including lower ADE, cost-effectiveness, simple facility requirements, scalability, and potential industrial-scale production in GMP facilities.

Oryza sativa L. is the world's most essential and economically important food crop. Climate change and ecological imbalances make rice plants vulnerable to abiotic and biotic stresses, threatening global food security. The Alfin-like (AL) transcription factor family plays a crucial role in plant development and stress responses. This study comprehensively analyzed this gene family and their expression profiles in rice, revealing nine AL genes, classifying them into three distinct groups based on phylogenetic analysis and identifying four segmental duplication events. RNA-seq data analysis revealed high expression levels of OsALs in different tissues, growth stages, and their responsiveness to stresses. RT-qPCR data showed significant expression of OsALs in different abiotic stresses. Identification of potential cis-regulatory elements in promoter regions has also unveiled their involvement. Tertiary structures of the proteins were predicted. These findings would lay the groundwork for future research to reveal their molecular mechanism in stress tolerance and plant development.

Novel anticancer drugs of natural origin have increased tremendously due to the resistance of multiple chemotherapeutic drugs in breast cancer therapy and their high toxicity with undesirable side effects. The study investigates the bioactivity of secondary metabolites derived from Bacillus cereus PSMS6 isolated from marine soil sediment in the Velar estuary, Parangepattai, Cuddalore district, Tamil Nadu, and India. Strains were isolated and antagonistic activity was screened using the agar well diffusion method. B. cereus PSMS6 exhibited potency, and its crude extract was tested for antioxidant, anticancer, and cytotoxic MTT assay potential. The methanolic extract of B. cereus PSMS6 was analyzed by mass spectrometry HRLC-MS and FT-IR to determine the bioactive compounds. A drug interaction study with the anti-breast cancer protein HER2 was performed by molecular docking analysis. Antioxidant activities were determined using total antioxidant scavenging assay, ABTS and DPPH free radical scavenging assays. The total antioxidant scavenging assay of the crude extract of B. cereus methanol had an IC₅₀ value of 28.33±1.01, in ABTS IC₅₀ value of the extract was 29.00±0.28 and in DPPH the IC₅₀ of the extract was 34.91±0.09. The negative ion compound Palmitoylglycerone phosphate had a LibDock score of 149.487 and the positive ion compound N5-(4-Methoxybenzyl) glutamine had 120.116. These compounds show promising anticancer activity. The current study reported that the bioactive secondary metabolite of B. cereus PSMS6 retains anti-cancer, and antioxidant properties. This is the first report to show the production of the Palmitoylglycerone phosphate metabolite from B. cereus PSMS6.

Cobra (Naja kaouthia) venom contains many toxins including α-neurotoxin (αNTX) and phospholipase A2 (PLA2), which can cause neurodegeneration, respiratory failure, and even death. The traditional antivenom derived from animal serum faces many challenges and limitations. Heavy-chain-only antibodies (HCAb), fusing V_HH with human IgG Fc region, offer advantages in tissue penetration, antigen binding, and extended half-life. This research involved the construction and transient expression of two types of V_HH-F_C which are specific to α-Neurotoxin (V_HH-αNTX-F_C) and Phospholipase A2 (V_HH-PLA2-F_C) in Nicotiana benthamiana leaves. The recombinant HCAbs were incubated for up to six days to optimize expression levels followed by purification by affinity chromatography and characterization using LC/Q-TOF mass spectrometry (MS). Purified proteins demonstrated over 92 % sequence coverage and an average mass of around 82 kDa with a high-mannose N-glycan profile. An antigen binding assay showed that the V_HH-αNTX-Fc has a greater ability to bind to crude venom than V_HH-PLA2-Fc.