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BACKGROUND: The spread of the COVID-19 disease is the result of an infection caused by the SARS-CoV2 virus. Four crucial proteins, spike [S], membrane [M], nucleocapsid [N], and envelope [E] in coronaviruses have been considered to a large extent. OBJECTIVE: This research aimed to express the recombinant protein of a multiepitope immunogen construct and evaluate the immunogenicity of the multiepitope vaccine that was previously designed as a candidate immunogenic against SARS-Cov-2. MATERIALS AND METHODS: Plasmid pET26b was transferred to the expression host E. coli BL21 [DE3] and the recombinant protein was expressed with IPTG induction. The recombinant protein was purified by Ni-NTA column affinity chromatography, and western blotting was used to confirm it. Finally, mice were immunized with recombinant protein in three doses. Then, the interaction of the 3D structure of the vaccine with the human neutralizing antibodies3D structures [7BWJ and 7K8N] antibody was evaluated by docking and molecular dynamics simulation. RESULTS: The optimized gene had a codon compatibility index of 0.96. The expression of the recombinant protein of the SARS-Cov-2 vaccine in an E. coli host led to the production of the recombinant protein with a weight of about 70 kDa with a concentration of 0.7 mg/ml. Immunization of mice with recombinant protein of SARS-Cov-2 vaccine-induced IgG serum antibody response. Statistical analysis showed that the antibody titer in comparison with the control sample has a significant difference, and the antibody titer was acceptable up to 1/256000 dilution. The simulation of vaccine binding with human antibodies by molecular dynamics showed that Root Mean Square Deviation [RMSD], Root Mean Square Fluctuation [RMSF], Radius of Gyration, and H-bond as well as van der Waals energies and electrostatic of Molecular mechanics Poisson- Boltzmann surface area [MM/PBSA] analysis have stable interaction. CONCLUSION: This recombinant protein can probably be used as an immunogen candidate for the development of vaccines against SARS-CoV2 in future research.
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Proteases, enzymes that hydrolyze peptide bonds, have various applications in medicine, clinical applications, and pharmaceutical development. They are used in cancer treatment, wound debridement, contact lens cleaning, prion degradation, biofilm removal, and fibrinolytic agents. Proteases are also crucial in cardiovascular disease treatment, emphasizing the need for safe, affordable, and effective fibrinolytic drugs. Proteolytic enzymes and protease biosensors are increasingly used in diagnostic and therapeutic applications. Advanced technologies, such as nanomaterials-based sensors, are being developed to enhance the sensitivity, specificity, and versatility of protease biosensors. These biosensors are becoming effective tools for disease detection due to their precision and rapidity. They can detect extracellular and intracellular proteases, as well as fluorescence-based methods for real-time and label-free detection of virus-related proteases. The active utilization of proteolytic enzymatic biosensors is expected to expand significantly in biomedical research, in-vitro model systems, and drug development. We focused on journal articles and books published in English between 1982 and 2024 for this study.
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Since 1997, highly pathogenic avian influenza viruses, such as H5N1, have been recognized as a possible pandemic hazard to men and the poultry business. The rapid rate of mutation of H5N1 viruses makes the whole process of designing vaccines extremely challenging. Here, we used an in silico approach to design a multi-epitope vaccine against H5N1 influenza A virus using hemagglutinin (HA) and neuraminidase (NA) antigens. B-cell epitopes, Cytotoxic T lymphocyte (CTL) and Helper T lymphocyte (HTL) were predicted via IEDB, NetMHC-4 and NetMHCII-2.3 respectively. Two adjuvants consisting of Human ß-defensin-3 (HßD-3) along with pan HLA DR-binding epitope (PADRE) have been chosen to induce more immune response. Linkers including KK, AAY, HEYGAEALERAG, GPGPGPG and double EAAAK were utilized to link epitopes and adjuvants. This construct encodes a protein having 350 amino acids and 38.46 kDa molecular weight. Antigenicity of ~ 1, the allergenicity of non-allergen, toxicity of negative and solubility of appropriate were confirmed through Vaxigen, AllerTOP, ToxDL and DeepSoluE, respectively. The 3D structure of H5N1 was refined and validated with a Z-Score of - 0.87 and an overall Ramachandran of 99.7%. Docking analysis showed H5N1 could interact with TLR7 (docking score of - 374.08 and by 4 hydrogen bonds) and TLR8 (docking score of - 414.39 and by 3 hydrogen bonds). Molecular dynamics simulations results showed RMSD and RMSF of 0.25 nm and 0.2 for H5N1-TLR7 as well as RMSD and RMSF of 0.45 nm and 0.4 for H5N1-TLR8 complexes, respectively. Molecular Mechanics Poisson-Boltzmann Surface Area (MM/PBSA) confirmed stability and continuity of interaction between H5N1-TLR7 with the total binding energy of - 29.97 kJ/mol and H5N1-TLR8 with the total binding energy of - 23.9 kJ/mol. Investigating immune response simulation predicted evidence of the ability to stimulate T and B cells of the immunity system that shows the merits of this H5N1 vaccine proposed candidate for clinical trials.
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Virus da Influenza A Subtipo H5N1 , Vacinas , Animais , Humanos , Virus da Influenza A Subtipo H5N1/genética , Epitopos de Linfócito T/genética , Receptor 7 Toll-Like , Receptor 8 Toll-Like , Epitopos de Linfócito B , Biologia Computacional/métodos , Simulação de Acoplamento Molecular , Vacinas de Subunidades Antigênicas/genéticaRESUMO
Microbial resistance against common antibiotics has become one of the most serious threats to human health. The increasing statistics on this problem show the necessity of finding a way to deal with it. In recent years, antimicrobial peptides with unique properties and the capability of targeting a wide range of pathogens, have been considered as a potential for replacing common antibiotics. A small chitin-binding protein with anticandidal activity was isolated from Moringa oleifera seeds by Neto and colleagues in 2017, which very much resembled antimicrobial peptides. In this study, the antimicrobial protein 'AF-DP' was identified and characterized. AF-DP was heterologously expressed, purified, and characterized, and its 3D structure was predicted. Six molecular dynamic simulations were performed to investigate how the protein interacts with Gram-negative inner and outer, Gram-positive, fungal, cancerous, and normal mammalian membranes. Also, its antimicrobial and anticancer activity was assessed in vitro via minimum inhibition concentration (MIC) and MTT assays, respectively. This protein with 111 amino acids and a total net charge (of 10.5) has been predicted to be mainly composed of alpha helix and random coils. Its MIC affecting the growth of Escherichia coli, Staphylococcus aureus, and Candida albicans was 30 µg/ml, 100 µg/ml, and 100 µg/ml, respectively; AF-DP showed anticancer activity against MCF-7 breast cancer cell line. Scanning electron microscopic analysis confirmed the creation of pores and scratches on the surface of the bacterial membrane. The results of this research show that AF-DP can be a candidate for the production of new drugs as an AMP with antimicrobial activity.Communicated by Ramaswamy H. Sarma.
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One of the major categories of industrial enzymes, proteases is crucial to the survival of living things. The purpose of this research was to newly thermostable protease from the thermophilum Geobacillus stearothermophilus. With the conserved catalytic tetrad, protease (Protease JJ) is closely related to the serine proteases from the subtilisin S8 peptidase, according to phylogenetic tree analysis. The tertiary structure of Protease JJ was predicted structurally using RoseTTAFold, and it is a sandwich structure overall. Homology modeling validation showed Protease JJ was modeled in X-ray's protein areas, and it has gained a favored Ramachandran graph regarding Phi/Psi angels. Protease JJ showed structure stability through Molecular dynamics simulation in the presence of Tween20 and Methanol in 1% concentration. Also, Protease JJ exhibited thermal stability at 60 to 90 °C so that amino acid exposure of Protease JJ was low and constant throughout the MD simulation. Docking results of Protease JJ with BSA and ßcasein were simulated via MD and it was found that Protease JJ could interact with both BSA and ßcasein strongly. MM/PBSA analysis showed Protease JJ may be involved via more amino acids with BSA as well as established more interaction hydrogen bonds. Overall, evidence suggests Protease JJ probably has merit for future experimental investigation as a thermostable protease.Communicated by Ramaswamy H. Sarma.
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The keratin-degrading bacterium Bacillus licheniformis secretes a keratinase with potential industrial interest. Here, the Keratinase gene was intracellularly expressed in Escherichia coli BL21(DE3) using pET-21b (+) vector. Phylogenetic tree analysis showed that KRLr1 is closely related to Bacillus licheniformis keratinase that belongs to the serine peptidase/subtilisin-like S8 family. Recombinant keratinase appeared on the SDS-PAGE gel with a band of about 38 kDa and was confirmed by western blotting. Expressed KRLr1 was purified by Ni-NTA affinity chromatography with a yield of 85.96% and then refolded. It was found that this enzyme has optimum activity at pH 6 and 37°C. PMSF inhibited the KRLr1 activity and Ca2+ and Mg2+ increased the KRLr1 activity. Using keratin 1% as the substrate, the thermodynamic values were determined as Km 14.54 mM, kcat 912.7 × 10-3 (S-1), and kcat/Km 62.77 (M-1 S-1). Feather digestion by recombinant enzyme using HPLC method, showed that the amino acids cysteine, phenylalanine, tyrosine and lysine had the highest amount compared to other amino acids obtained from digestion. Molecular dynamics (MD) simulation of HADDOCK docking results exhibited that KRLr1 enzyme was able to interact strongly with chicken feather keratine 4 (FK4) compared to chicken feather keratine 12 (FK12). These properties make keratinase KRLr1 a potential candidate for various biotechnological applications.
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Viral hepatitis, the most common cause of inflammatory liver disease, affects hundreds of millions of people worldwide. It is most commonly associated with one of the five nominal hepatitis viruses (hepatitis A-E viruses). HBV and HCV can cause acute infections and lifelong, persistent chronic infections, while HAV and HEV cause self-limiting acute infections. HAV and HEV are predominantly transmitted through the fecal-oral route, while diseases transmitted by the other forms are blood-borne diseases. Despite the success in the treatment of viral hepatitis and the development of HAV and HBV vaccines, there is still no accurate diagnosis at the genetic level for these diseases. Timely diagnosis of viral hepatitis is a prerequisite for efficient therapeutic intervention. Due to the specificity and sensitivity of clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated sequences (Cas) technology, it has the potential to meet critical needs in the field of diagnosis of viral diseases and can be used in versatile point-of-care (POC) diagnostic applications to detect viruses with both DNA and RNA genomes. In this review, we discuss recent advances in CRISPR-Cas diagnostics tools and assess their potential and prospects in rapid and effective strategies for the diagnosis and control of viral hepatitis infection.
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Vírus da Hepatite A , Hepatite A , Viroses , Humanos , Sistemas CRISPR-Cas , Fezes , Infecção PersistenteRESUMO
Currently, there are two vaccines based on killed and/or weakened Salmonella bacteria, but no recombinant vaccine is available for preventing or treating the disease. We used an in silico approach to design a multi-epitope vaccine against Salmonella using OmpA, OmpS, SopB, SseB, SthA and FilC antigens. We predicted helper T lymphocyte, cytotoxic T lymphocyte, and IFN-γ epitopes. The FilC sequence was used as a bovine TLR5 agonist, and the linkers KK, AAY, GPGPG and EAAAK were used to connect epitopes. The final sequence consisted of 747 amino acid residues, and the expressed soluble protein (â¼79.6 kDa) was predicted to be both non-allergenic and antigenic. The tertiary structure of modeled protein was refined and validated, and the interactions of vaccine 3D structure were evaluated using molecular docking, and molecular dynamics simulation (RMSD, RMSF and Gyration). This structurally stable protein could interact with human TLR5. The C-ImmSim server predicted that this proposed vaccine likely induces an immune response by stimulating T and B cells, making it a potential candidate for further evaluation for the prevention and treatment of Salmonella infection.
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Receptor 5 Toll-Like , Fatores de Virulência , Animais , Bovinos , Humanos , Simulação de Acoplamento Molecular , Epitopos de Linfócito T , Epitopos de Linfócito B , Vacinas de Subunidades Antigênicas , Salmonella/genética , Biologia ComputacionalRESUMO
BACKGROUND: Contrary to the advantageous anticancer activities of curcumin (Cur), limited bioavailability and solubility hindered its efficacy. Here, nontoxic dendrosomal nano carrier with Cur was used to overcome these problems. Despite considerable antitumor properties of Oxaliplatin (Oxa), the limiting factors are drug resistance and adverse side-effects. The hypothesis of this study was to evaluate the possible synergism between dendrosomal nanocurcumin (DNC) and Oxa and these agents showed growth regulatory effects on SKOV3 and OVCAR3 cells. METHODS AND MATERIALS: In the present study, colony formation, wound healing motility, cell adhesion, transwell invasion and migration assay and cell cycle arrest with or without DNC, Oxa and Combination were defined. In addition to, real time PCR and Western blot were used to analyze AKT, PI3K, PKC, JNK, P38 and MMPs mRNAs and proteins expressions. Docking of MMP-2-Cur, MMP-2-DNC and MMP-2-Oxa was performed and the results of all three complexes were simulated by molecular dynamics. RESULTS: Our findings illustrated that DNC had the greatest effect on cell death as compared to the Cur alone. Moreover, the growth inhibitory effects (such as cell death correlated to apoptosis) were more intense if Oxa was added followed by DNC at 4 h interval. However, insignificant effects were observed upon simultaneous addition of these two agents in both cell lines. Besides, a combination of agents synergistically alters the relative expression of MMP-9. CONCLUSIONS: The docking results showed that His70 and Asp100 may play a key role at the MMP-2 binding site. The matrigel invasion as well as cell viability of ovarian cancer cell lines SKOV3 and OVCAR3 by DNC alone or in combination with Oxa was inhibited significantly. The inhibitory effects of these agents were due to the differential expression levels of MMP 2 and MMP 9 regulated by multiple downstream signaling cascades. From the molecular dynamic simulation studies, it was confirmed that DNC established a strong interaction with MMP-2.
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Curcumina , Neoplasias Ovarianas , Humanos , Feminino , Oxaliplatina/farmacologia , Apoptose , Metaloproteinase 2 da Matriz/farmacologia , Neoplasias Ovarianas/tratamento farmacológico , Linhagem Celular Tumoral , Proliferação de Células , Curcumina/farmacologia , Movimento CelularRESUMO
BACKGROUND: Contrary to the advantageous anticancer activities of curcumin (Cur), limited bioavailability and solubility hindered its efficacy. Here, nontoxic dendrosomal nano carrier with Cur was used to overcome these problems. Despite considerable antitumor properties of Oxaliplatin (Oxa), the limiting factors are drug resistance and adverse side-effects. The hypothesis of this study was to evaluate the possible synergism between dendrosomal nanocurcumin (DNC) and Oxa and these agents showed growth regulatory effects on SKOV3 and OVCAR3 cells. METHODS: and materials In the present study, colony formation, wound healing motility, cell adhesion, transwell invasion and migration assay and cell cycle arrest with or without DNC, Oxa and Combination were defined. In addition to, real time PCR and Western blot were used to analyze AKT, PI3K, PKC, JNK, P38 and MMPs mRNAs and proteins expressions. Docking of MMP-2-Cur, MMP-2-DNC and MMP-2-Oxa was performed and the results of all three complexes were simulated by molecular dynamics. RESULTS: Our findings illustrated that DNC had the greatest effect on cell death as compared to the Cur alone. Moreover, the growth inhibitory effects (such as cell death correlated to apoptosis) were more intense if Oxa was added followed by DNC at 4 h interval. However, insignificant effects were observed upon simultaneous addition of these two agents in both cell lines. Besides, a combination of agents synergistically alters the relative expression of MMP-9. CONCLUSIONS: The docking results showed that His70 and Asp100 may play a key role at the MMP-2 binding site. The matrigel invasion as well as cell viability of ovarian cancer cell lines SKOV3 and OVCAR3 by DNC alone or in combination with Oxa was inhibited significantly. The inhibitory effects of these agents were due to the differential expression levels of MMP 2 and MMP 9 regulated by multiple downstream signaling cascades. From the molecular dynamic simulation studies, it was confirmed that DNC established a strong interaction with MMP-2.
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Humanos , Feminino , Neoplasias Ovarianas/tratamento farmacológico , Curcumina/farmacologia , Movimento Celular , Apoptose , Metaloproteinase 2 da Matriz/farmacologia , Linhagem Celular Tumoral , Proliferação de Células , Oxaliplatina/farmacologiaRESUMO
Vaccines against Brucella abortus, B. melitensis and B. suis have been based on weakened or killed bacteria, however there is no recombinant vaccine for disease prevention or therapy. This study attempted to predict IFN-γ epitopes, T cell cytotoxicity, and T lymphocytes in order to produce a multiepitope vaccine based on BtpA, Omp16, Omp28, virB10, Omp25, and Omp31 antigens against B. melitensis, B. abortus, and B. suis. AAY, GPGPG, and EAAAK peptides were used as epitope linkers, while the PADRE sequence was used as a Toll-like receptor 2 (TLR2) and TLR4 agonist. The final construct included 389 amino acids, and was a soluble protein with a molecular weight of 41.3 kDa, and nonallergenic and antigenic properties. Based on molecular docking studies, molecular dynamics simulations such as Gyration, RMSF, and RMSD, as well as tertiary structure validation methods, the modeled protein had a stable structure capable of interacting with TLR2/4. As a result, this novel vaccine may stimulate immune responses in B and T cells, and could prevent infection by B. suis, B. abortus, and B. melitensis.
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Brucella melitensis , Brucelose , Humanos , Receptor 2 Toll-Like , Mapeamento de Epitopos , Epitopos de Linfócito T , Brucelose/prevenção & controle , Brucelose/microbiologia , Receptor 4 Toll-Like , Simulação de Acoplamento Molecular , Antígenos de Bactérias/química , AminoácidosRESUMO
Cellulases are hydrolytic enzymes with wide scientific and industrial applications. We described a novel cellulase, CelC307, from the thermophilic indigenous Cohnella sp. A01. The 3-D structure of the CelC307 was predicted by comparative modeling. Docking of CelC307 with specific inhibitors and molecular dynamic (MD) simulation revealed that these ligands bound in a non-competitive manner. The CelC307 protein was purified and characterized after recombinant expression in Escherichia coli (E. coli) BL21. Using CMC 1% as the substrate, the thermodynamic values were determined as Km 0.46 mM, kcat 104.30 × 10-3 (S-1), and kcat/Km 226.73 (M-1 S-1). The CelC307 was optimally active at 40 °C and pH 7.0. The culture condition was optimized for improved CelC307 expression using Plackett-Burman and Box-Behnken design as follows: temperature 20 °C, pH 7.5, and inoculation concentration with an OD600 = 1. The endoglucanase activity was positively modulated in the presence of Na+, Li+, Ca2+, 2-mercaptoethanol (2-ME), and glycerol. The thermodynamic parameters calculated for CelC307 confirmed its inherent thermostability. The characterized CelC307 may be a suitable candidate for various biotechnological applications.
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Bacillales , Celulase , Celulases , Bacillales/metabolismo , Celulase/metabolismo , Celulases/metabolismo , Estabilidade Enzimática , Escherichia coli/genética , Escherichia coli/metabolismo , Concentração de Íons de Hidrogênio , Íons , TemperaturaRESUMO
A new strain of Influenza A Virus (IAV), so-called "H7N9 Avian Influenza", is the first strain of this virus in which a human is infected by transmitting the N9 of influenza virus. Although continuous human-to-human transmission has not been reported, the occurrence of various H7N9-associated epidemics and the lack of production of strong antibodies against H7N9 in humans warn of the potential for H7N9 to become a new pandemic. Therefore, the need for effective vaccination against H7N9 as a life-threatening viral pathogen has become a major concern. The current study reports the design of a multi-epitope vaccine against Hemagglutinin (HA) and Neuraminidase (NA) proteins of H7N9 Influenza A virus by prediction of Cytotoxic T lymphocyte (CTL), Helper T lymphocyte (HTL), IFN-γ and B-cell epitopes. Human ß-defensin-3 (HßD-3) and pan HLA DR-binding epitope (PADRE) sequence were considered as adjuvant. EAAAK, AAY, GPGPG, HEYGAEALERAG, KK and RVRR linkers were used as a connector for epitopes. The final construct contained 777 amino acids that are expected to be a recombinant protein of about ~ 86.38 kDa with antigenic and non-allergenic properties after expression. Modeled protein analysis based on the tertiary structure validation, docking studies, and molecular dynamics simulations results like Root-mean-square deviation (RMSD), Gyration, Root-mean-square fluctuation (RMSF) and Molecular Mechanics Poisson-Boltzmann Surface Area (MM/PBSA) showed that this protein has a stable construct and capable of being in interaction with Toll-like receptor 7 (TLR7), TLR8 and m826 antibody. Analysis of the obtained data the demonstrates that suggested vaccine has the potential to induce the immune response by stimulating T and Bcells, and may be utilizable for prevention purposes against Avian Influenza A (H7N9).
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Epitopos/imunologia , Subtipo H7N9 do Vírus da Influenza A/imunologia , Vacinas contra Influenza/imunologia , Influenza Humana/imunologia , Animais , Aves , Biologia Computacional , Simulação por Computador , Glicoproteínas de Hemaglutininação de Vírus da Influenza/imunologia , Humanos , Imunidade , Influenza Aviária/imunologia , Modelos Imunológicos , Modelos Moleculares , Neuraminidase/imunologia , Proteínas Virais/imunologiaRESUMO
Therapeutic vaccines are an effective approach in cancer therapy for treating the disease at later stages. The Food and Drug Administration (FDA) recently approved the first therapeutic cancer vaccine, and further studies are ongoing in clinical trials. These are expected to result in the future development of vaccines with relatively improved efficacy. Several vaccination approaches are being studied in pre-clinical and clinical trials, including the generation of anti-cancer vaccines by plant expression systems.This approach has advantages, such as high safety and low costs, especially for the synthesis of recombinant proteins. Nevertheless, the development of anti-cancer vaccines in plants is faced with some technical obstacles.Herein, we summarize some vaccines that have been used in cancer therapy, with an emphasis on plant-based vaccines.
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Vacinas Anticâncer/uso terapêutico , Neoplasias/terapia , Plantas Geneticamente Modificadas/genética , Animais , HumanosRESUMO
The agricultural sector is currently facing many global challenges, such as climate change, and environmental problems such as the release of pesticides and fertilizers, which will be exacerbated in the face of population growth and food shortages. Therefore, the need to change traditional farming methods and replace them with new technologies is essential, and the application of nanotechnology, especially green technology offers considerable promise in alleviating these problems. Nanotechnology has led to changes and advances in many technologies and has the potential to transform various fields of the agricultural sector, including biosensors, pesticides, fertilizers, food packaging and other areas of the agricultural industry. Due to their unique properties, nanomaterials are considered as suitable carriers for stabilizing fertilizers and pesticides, as well as facilitating controlled nutrient transfer and increasing crop protection. The production of nanoparticles by physical and chemical methods requires the use of hazardous materials, advanced equipment, and has a negative impact on the environment. Thus, over the last decade, research activities in the context of nanotechnology have shifted towards environmentally friendly and economically viable 'green' synthesis to support the increasing use of nanoparticles in various industries. Green synthesis, as part of bio-inspired protocols, provides reliable and sustainable methods for the biosynthesis of nanoparticles by a wide range of microorganisms rather than current synthetic processes. Therefore, this field is developing rapidly and new methods in this field are constantly being invented to improve the properties of nanoparticles. In this review, we consider the latest advances and innovations in the production of metal nanoparticles using green synthesis by different groups of microorganisms and the application of these nanoparticles in various agricultural sectors to achieve food security, improve crop production and reduce the use of pesticides. In addition, the mechanism of synthesis of metal nanoparticles by different microorganisms and their advantages and disadvantages compared to other common methods are presented.
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Agricultura/métodos , Nanopartículas Metálicas , Nanotecnologia/métodos , Técnicas Biossensoriais , Proteção de Cultivos , Fertilizantes , Segurança Alimentar , Fungicidas Industriais , Química Verde , Nanoestruturas , PraguicidasRESUMO
Probiotics are beneficial bacteria that exist within the human gut, and which are also present in different food products and supplements. They have been investigated for some decades, due to their potential beneficial impact on human health. Probiotics compete with pathogenic microorganisms for adhesion sites within the gut, to antagonize them or to regulate the host immune response resulting in preventive and therapeutic effects. Therefore, dysbiosis, defined as an impairment in the gut microbiota, could play a role in various pathological conditions, such as lactose intolerance, gastrointestinal and urogenital infections, various cancers, cystic fibrosis, allergies, inflammatory bowel disease, and can also be caused by antibiotic side effects. MicroRNAs (miRNAs) are short non-coding RNAs that can regulate gene expression in a post-transcriptional manner. miRNAs are biochemical biomarkers that play an important role in almost all cellular signaling pathways in many healthy and disease states. For the first time, the present review summarizes current evidence suggesting that the beneficial properties of probiotics could be explained based on the pivotal role of miRNAs. Video Abstract.
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MicroRNAs , Probióticos/uso terapêutico , Animais , HumanosRESUMO
Chitosan nanomaterials have become a hot topic in biomedicine due to exerting antimicrobial effects with interestingly high levels of biodegradability and biocompatibility without causing toxicity. Regarded as a potential means of wound dressing with antimicrobial activity, chitosan exhibits higher efficiency when it is functionally modified with other natural compounds, metallic antimicrobial particles and antibiotics. Mechanistically, the antibacterial effect of chitosan is mostly, associated with the death-proceeding leakage of intracellular content, induced by malfunction and altered permeability of the negatively charged cell membrane, on which chitosan is adsorbed. Moreover, chitosan nanoparticles (NPs) are endowed with favorable features of NPs (i.e., large surface-to-volume ratio, high functionalization possibilities and a greater capacity for drug loading), as well as that of their chitosan base, thereby possessing strengthened antibacterial potential. In addition, polycations target negatively charged bacterial membranes, so bacteria cells are more strongly affected by polycationic chitosan NPs than pure chitosan.
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Antibacterianos/farmacologia , Infecções Bacterianas/tratamento farmacológico , Quitosana/administração & dosagem , Nanopartículas/administração & dosagem , Animais , Antibacterianos/química , Quitosana/química , Humanos , Nanopartículas/químicaRESUMO
Proteases play an essential role in living organisms and represent one of the largest groups of industrial enzymes. The aim of this work was recombinant production and characterization of a newly identified thermostable protease 1147 from thermophilum indigenous Cohnella sp. A01. Phylogenetic tree analysis showed that protease 1147 is closely related to the cysteine proteases from DJ-1/ThiJ/PfpI superfamily, with the conserved catalytic tetrad. Structural prediction using MODELLER 9v7 indicated that protease 1147 has an overall α/ß sandwich tertiary structure. The gene of protease 1147 was cloned and expressed in Escherichia coli (E. coli) BL21. The recombinant protease 1147 appeared as a homogenous band of 18 kDa in SDS-PAGE, which was verified by western blot and zymography. The recombinant protein was purified with a yield of approximately 88% in a single step using Ni-NTA affinity chromatography. Furthermore, a rapid one-step thermal shock procedure was successfully implemented to purify the protein with a yield of 73%. Using casein as the substrate, Km, and kcat, kcat/Km values of 13.72 mM, 3.143 × 10-3 (s-1), and 0.381 (M-1 S-1) were obtained, respectively. The maximum protease activity was detected at pH = 7 and 60°C with the inactivation rate constant (kin) of 2.10 × 10-3 (m-1), and half-life (t1/2) of 330.07 min. Protease 1147 exhibited excellent stability to organic solvent, metal ions, and 1% SDS. The protease activity was significantly enhanced by Tween 20 and Tween 80 and suppressed by cysteine protease specific inhibitors. Docking results and molecular dynamics (MD) simulation revealed that Tween 20 interacted with protease 1147 via hydrogen bonds and made the structure more stable. CD and fluorescence spectra indicated structural changes taking place at 100°C, very basic and acidic pH, and in the presence of Tween 20. These properties make this newly characterized protease a potential candidate for various biotechnological applications.
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Bacillales/enzimologia , Proteínas de Bactérias/química , Peptídeo Hidrolases/química , Proteínas de Bactérias/isolamento & purificação , Proteínas de Bactérias/ultraestrutura , Clonagem Molecular , Ensaios Enzimáticos , Estabilidade Enzimática , Concentração de Íons de Hidrogênio , Simulação de Dinâmica Molecular , Peso Molecular , Peptídeo Hidrolases/isolamento & purificação , Peptídeo Hidrolases/ultraestrutura , Proteínas Recombinantes/química , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/ultraestrutura , Especificidade por SubstratoRESUMO
Both environmental and genetic factors are involved in the initiation and development of gastrointestinal cancer. Covalent closed circular RNAs (circRNAs) are produced by a mechanism called "back-splicing" from mRNAs. They are highly stable and show cell and tissue specific expression patterns. Although some functions such as "microRNA sponge" and "RNA binding protein sponge" have been reported for a small number of circRNAs, the function of thousands of other circRNAs is still unknown. Dysregulation of circRNAs has been reported in many GI cancers and are involved in metastasis and invasion. CircRNAs have been reported to be useful as prognostic markers and targets for developing new treatments. We first describe the properties and biogenesis of circRNAs. We then summarize recent reports about circRNA functions, expression status, and their potential to be used as biomarkers in GI cancers including, gastric cancer, colorectal cancer, esophageal cancer, hepatocellular carcinoma, gallbladder cancer and pancreatic cancer.