Binding Domain Protein Research Articles

Exploring the antimicrobial and antioxidative potential of Leucas aspera (Willd.) Link: Phytochemical screening, molecular docking, and HR-LC/MS profiling against SARS-CoV-2 protein 3CLPro, Spike and RDB

BackgroundLeucas aspera (Willd.) Link, a member of the Lamiaceae family and commonly known as "Thumbai," is found throughout India and has been utilized in traditional Indian medicine to treat various ailments. PurposeThis study entails methanolic extraction of Leucas aspera (Willd.) Link using a Soxhlet apparatus, followed by phytochemical screening and antioxidant activity assessment. MethodsThe L. aspera methanolic leaf extract displayed a phenolic content of 100 µg/ml, consisting of 3.02 mg of gallic acid dry weight equivalents and a flavonoid content of 100 µg/ml with 3.35 mg quercetin equivalents per dry weight. The extract's free radical scavenging capabilities were at 150 µg/ml in DPPH, showing an 87% effectiveness compared to the standard ascorbic acid 49% inhibition capacity. The ABTS radical scavenging activity in 100 µg/ml extract measured 64.32%, phosphomolybdate assay indicated 86.89%, and hydroxide radical scavenging capacity showed 126.72%. Antibacterial activity was also assessed through agar well diffusion assays against Klebsiella pneumoniae, Salmonella typhi, Staphylococcus aureus, and Leucobacter sp., Where K. pneumoniae exhibits the highest zone of inhibition. The methanolic crude extract of L. aspera was further using TLC, FT-IR and HR-LC/MS to identify functional groups and phytocompound present in the extract. ResultsHRLC-MS was used to conduct metabolite profiling of the methanolic crude extract of L. aspera leaves, discovering 37 positive and 43 negative compounds. To evaluate the ADMET properties and predict the drug-likeness of these 80 phytochemicals, an in silico analysis was performed. The results of this analysis revealed that only 19 of the compounds met the ADMET limitations and had suitable Log P values. Molecular docking of the 19 compounds against the SARS-CoV-2 main protease (3CLPro) protein (PDB ID: 6LU7) and spike protein receptor binding domain (SGp-RBD) protein (PDB ID: 2GHV) revealed Famprofazone and Loxtidine compounds having the highest binding affinity with LibDock scores of 105.53 and 116.70 respectively. ConclusionThus, our findings could serve as potential active molecules of L. aspera against this target protein. This study provides further proof of bioactive compounds produced by the L. aspera leaf extract.

Transcription factors in tanshinones: Emerging mechanisms of transcriptional regulation

Transcription factors play a crucial role in the biosynthesis of tanshinones, which are significant secondary metabolites derived from Salvia miltiorrhiza, commonly known as Danshen. These compounds have extensive pharmacological properties, including anti-inflammatory and cardioprotective effects. This review delves into the roles of various transcription factor families, such as APETALA2/ethylene response factor, basic helix-loop-helix, myeloblastosis, basic leucine zipper, and WRKY domain-binding protein, in regulating the biosynthetic pathways of tanshinones. We discuss the emerging mechanisms by which these transcription factors influence the synthesis of tanshinones, both positively and negatively, by directly regulating gene expression or forming complex regulatory networks. Additionally, the review highlights the potential applications of these insights in enhancing tanshinone production through genetic and metabolic engineering, setting the stage for future advancements in medicinal plant research.

The KLF16/MYC feedback loop is a therapeutic target in bladder cancer

BackgroundBladder cancer (BLCA) is a common malignancy characterized by dysregulated transcription and a lack of effective therapeutic targets. In this study, we aimed to identify and evaluate novel targets with clinical potential essential for tumor growth in BLCA.MethodsCRISPR-Cas9 screening was used to identify transcription factors essential for bladder cancer cell viability. The biological functions of KLF16 in bladder cancer were investigated both in vitro and in vivo. The regulatory mechanism between KLF16 and MYC was elucidated through a series of analyses, including RNA sequencing, quantitative polymerase chain reaction (qPCR), RNA immunoprecipitation, Western blotting, Mass spectrometry, Dual-luciferase reporter assays, Cleavage Under Targets and Tagmentation (CUT&Tag) sequencing, OptoDroplets assays, and RNA stability assay. The clinical relevance of KLF16 and MYC in bladder cancer was evaluated through analyses of public databases and immunohistochemistry.ResultsKrüppel-like factor 16 (KLF16) was essential for BLCA cell viability. Elevated expression of KLF16 was observed in bladder cancer tissues, and higher expression levels of KLF16 were correlated with poor progression-free survival (PFS) and cancer-specific survival (CSS) probabilities in BLCA patients. Mechanistically, KLF16 mRNA competed with the mRNA of dual-specificity phosphatase 16 (DUSP16) for binding to the RNA-binding protein, WW domain binding protein 11 (WBP11), resulting in destabilization of the DUSP16 mRNA. This, in turn, led to activation of ERK1/2, which stabilized the MYC protein. Furthermore, KLF16 interacted with MYC to form nuclear condensates, thereby enhancing MYC’s transcriptional activity. Additionally, MYC transcriptionally upregulated KLF16, creating a positive feedback loop between KLF16 and MYC that amplified their oncogenic functions. Targeting this loop with bromodomain inhibitors, such as OTX015 and ABBV-744, suppressed the transcription of both KLF16 and MYC, resulting in reduced BLCA cell viability and tumor growth, as well as increased sensitivity to chemotherapy.ConclusionsOur study revealed the crucial role of the KLF16/MYC regulatory axis in modulating tumor growth and chemotherapy sensitivity in BLCA, suggesting that combining bromodomain inhibitors, such as OTX015 or ABBV-744, with DDP or gemcitabine could be a promising therapeutic intervention for BLCA patients.

Leveraging Walnut Somatic Embryos as a Biomanufacturing Platform for Recombinant Proteins and Metabolites.

Biomanufacturing enables novel sources of compounds with constant demand, such as food coloring and preservatives, as well as new compounds with peak demand, such as diagnostics and vaccines. The COVID-19 pandemic has highlighted the need for alternative sources of research materials, thrusting research on diversification of biomanufacturing platforms. Here, we show initial results exploring the walnut somatic embryogenic system expressing the recombinant receptor binding domain (RBD) and ectodomain of the spike protein (Spike) from the SARS-CoV-2 virus. Stably transformed walnut embryo lines were selected and propagated in vitro. Both recombinant proteins were detected at 3-14 µg/g dry weight of tissue culture material. Although higher yields of recombinant protein have been obtained using more conventional biomanufacturing platforms, we also report on the production of the red pigment betanin in somatic embryos, reaching yields of 650 mg/g, even higher than red beet Beta vulgaris. This first iteration shows the potential of biomanufacturing using somatic walnut embryos that can now be further optimized for different applications sourcing specialized proteins and metabolites.

Silencing of SH3BP2 Inhibits Microglia Activation Via the JAK/STAT Signaling in Spinal Cord Injury Models.

The purpose of our study was to investigate the expression of SH3 domain-binding protein 2 (SH3BP2) in spinal cord injury (SCI) rats and lipopolysaccharide (LPS)-induced microglia, and explored its impact as well as potential mechanism. We examined the level of SH3BP2 in SCI rats using GEO data, immunofluorescence co-staining, qRT-PCR and western blotting. Next, we constructed a rat model with SH3BP2 silencing by injecting LV-shSH3BP2 into the injury site of SCI rats, and then evaluated its neurological outcome, functional recovery, M1 polarization and neuroinflammation by Basso-Beattie-Bresnahan (BBB) score, inclined plane test, Nissl staining and hematoxylin-eosin (H&E). The SH3BP2-related signaling pathway was predicted by KEGG analysis in GSE45006 dataset. BV2 microglial cells and primary microglia were incubated with LPS, and then measured its activation and inflammation by qRT-PCR, western blotting and immunofluorescence. Further complement experiments were performed to explore the molecular mechanisms of SH3BP2. The expression of SH3BP2 was increased in the spinal dorsal horn tissues of SCI rats and LPS-induced microglia. Silencing of SH3BP2 improved neurological outcomes and functional recovery, attenuated neuroinflammation and microglia polarization in SCI rats. Additionally, the JAK/STAT pathway was regulated by SH3BP2. Silencing of SH3BP2 inhibited LPS-induced microglia inflammation and activation, decreased the phosphorylation levels of JAK and STAT. Silencing of SH3BP2 attenuated SCI by regulating the JAK/STAT pathway to inhibit the activation of microglia.

In vivo characterization of ACE2 expression in Sprague-Dawley rats and cultured primary brain pericytes highlights the utility of Rattus norvegicus in the study of COVID-19 brain pathophysiology

A high number of COVID-19 patients report ongoing neurological impairments including headache, fatigue and memory impairments. Our understanding of COVID-19 disease mechanisms in the brain is limited and relies on post-mortem human tissues, in vitro studies in various cell lines (both human and animal) as well as preclinical studies in a variety of species. Notably the use of rats in the study of COVID-19 has been scarce in part due to early reports of low infectivity of the original Wuhan strain in mice and rats. Evidence has shown that subsequent strains that have mutated from the original strain are capable of infection in rats. Here we present an immunohistological characterization of ACE2 expression in the rat brain perivascular region. We found ACE2 to be expressed in pericytes but not endothelial cells or astrocytes in the perivascular space. We further examined the uptake of Omicron variants 1.1.529 and BA.2 receptor binding domains (RBD) of the SARS-CoV2 spike protein in primary brain pericytes derived from rats. We demonstrate that rat primary brain pericytes are susceptible to SARS-CoV2 spike protein uptake and induce functional changes in pericytes associated with a reduction in tight junction protein expression. These data provide evidence that rat primary cell responses to SARS-CoV2 infection are consistent with reports of infectivity in other species (transgenic mice expressing hACE2, ferrets, hamsters) and supports the use of this model organism with a long history of use in the study of disease which should be leveraged for study of COVID-19 in the brain.

Adaptor protein 3BP2 regulates gene expression in addition to the ubiquitination and proteolytic activity of MALT1 in dectin-1-stimulated cells.

Dectin-1, a C-type lectin, plays important roles in the induction of antifungal immunity. Caspase recruitment domain-containing protein 9 (CARD9) is essential for the dectin-1-induced production of cytokines through the activation of NF-κB. However, the molecular mechanisms underlying the dectin-1-mediated activation of CARD9 have not been fully elucidated. Recently, we reported that the adaptor protein SH3 domain-binding protein 2 (3BP2) is required for the dectin-1-induced production of cytokines and activation of NF-κB, although the relationship between 3BP2 and CARD9 in dectin-1-mediated signaling remains unclear. Here, we report that 3BP2 is required for dectin-1-induced expression of several genes that may contribute to antifungal immunity in bone marrow-derived dendritic cells (BMDCs). The results of reporter assays using HEK-293T cells indicate that 3BP2 induces CARD9-mediated activation of NF-κB through B-cell leukemia/lymphoma 10, mucosa-associated lymphoid tissue lymphoma translocation protein 1 (MALT1), and TNF receptor-associated factor 6-dependent mechanisms. In addition, we show that 3BP2 induces CARD9-mediated ubiquitination of cellular proteins and that MALT1 cleaves 3BP2 in a CARD9-dependent manner. Furthermore, we show that 3BP2 is required for the ubiquitination, in addition to the activation, of MALT1, which leads to MALT1-depenedent cleavage of 3BP2 in dectin-1-stimulated BMDCs. Finally, we identified hematopoietic cell-specific Lyn substrate 1 as a target of 3BP2, which is essential for dectin-1-induced expression of interleukin 10 in BMDCs. These results indicate that 3BP2 regulates gene expression and functions of MALT1 in dectin-1-stimulated cells and that 3BP2 plays an important role in the dectin-1-mediated antifungal immunity.

Response of Foldable Protein Conformations to Non-Physiological Perturbations: Interplay of Thermal Factors and Confinement.

Technological advances frequently interface biomolecules with nanomaterials at non-physiological conditions, necessitating response characterization of key processes. Similar encounters are expected in cellular contexts. We report in silico investigations of the response of diverse protein conformational states to lowering of temperature and imposition of spatial constraints. Conformational states are represented by folded form of the Albumin binding domain (ABD) protein, its compact denatured form, and structurally disordered nascent folding elements. Data from extensive simulations are evaluated to elicit structural, thermodynamic and dynamic responses of the states and their associated environment. Analyses reveal alterations to folding propensity with reduced thermal energy and confinement, with signatures of trend reversal in highly disordered states. Across temperatures, confinement has restrictive effects on volume and energetic fluctuations, leading to narrowing of differences in isothermal compressibility (κ) and heat capacities (Cp). While excess (over ideal gas) entropy of the hydration layer marks dependence on the conformational state at bulk, confinement triggers erasure of differences. These observations are largely consistent with timescales of protein-water hydrogen bonding dynamics. The results implicate multi-factorial associations within a simple bio-nano complex. We expect the current study to motivate investigations of more biologically relevant interfaces towards mechanistic understanding and potential applications.

Chemical Proteomics Approaches for Screening Small Molecule Inhibitors Covalently Binding to SARS-Cov-2.

Although various strategies have been used to prevent and treat SARS-CoV-2, the spread and evolution of SARS-CoV-2 is still progressing rapidly. The emerging variants Omicron and its sublineage have a greater ability to spread and escape nearly all current monoclonal antibodies treatments, highlighting an urgent need to develop therapeutics targeting current and emerging Omicron variants or recombinants with breadth and potency. Here, some small molecule drugs are rapidly identified that could covalently binding to receptor binding domain (RBD) protein of Omicron through the combined application of artificial intelligence (AI) and activity-based protein profiling (ABPP) technology. The surface plasmon resonance (SPR) and pseudo-virus neutralization experiments further reveal that an FDA-approved drug gallic acid has robust neutralization potency against Omicron pseudo-virus with the IC50 values of 23.56 × 10-6 m. Taken together, a platform combining AI intelligence, biochemical, SPR, molecular docking, and pseudo-virus-based screening for rapid identification and evaluation of potential anti-SARS-CoV-2 small molecule drugs is established and the effectiveness of the platform is validated.

Role of Circular RNA MMP9 in Glioblastoma Progression: From Interaction With hnRNPC and hnRNPA1 to Affecting the Expression of BIRC5 by Sequestering miR-149.

Glioblastoma multiforme (GBM) presents a significant challenge in neuro-oncology due to its aggressive behavior and self-renewal capacity. Circular RNAs (circRNAs), a subset of non-coding RNAs (ncRNAs) generated through mRNA back-splicing, are gaining attention as potential targets for GBM research. In our study, we sought to explore the functional role of circMMP9 (circular form of matrix metalloproteinase-9) as a promising therapeutic target for GBM through bioinformatic predictions and human data analysis. Our results suggest that circMMP9 functions as a sponge for miR-149 and miR-542, both upregulated in GBM based on microarray data. Kaplan-Meier analysis indicated that reduced levels of miR-149 and miR-542 correlate with worse survival outcomes in GBM, suggesting their role as tumor suppressors. Importantly, miR-149 has been demonstrated to inhibit the expression of BIRC5 (baculoviral inhibitor of apoptosis repeat-containing 5 or survivin), a significant promoter of proliferation in GBM. BIRC5 is not only upregulated in GBM but also in various other cancers, including neuroblastoma and other brain cancers. Our protein-protein interaction analysis highlights the significance of BIRC5 as a central hub gene in GBM. CircMMP9 seems to influence this complex relationship by suppressing miR-149 and miR-542, despite their increased expression in GBM. Additionally, we found that circMMP9 directly interacts with heterogeneous nuclear ribonucleoproteins C and A1 (hnRNPC and A1), although not within their protein-binding domains. This suggests that hnRNPC/A1 may play a role in transporting circMMP9. Moreover, RNA-seq data from GBM patient samples confirmed the increased expression of BIRC5, PIK3CB, and hnRNPC/A1, further emphasizing the potential therapeutic significance of circMMP9 in GBM. In this study, we propose for the first time a new epigenetic regulatory role for circMMP9, highlighting a novel aspect of its oncogenic function. circMMP9 may regulate BIRC5 expression in GBM by sponging miR-149 and miR-542. BIRC5, in turn, suppresses apoptosis and enhances proliferation in GBM. Nonetheless, more extensive studies are advised to delve deeper into the roles of circMMP9, especially in the context of glioma.

Targeting Human Papillomavirus 33 E2 DNA Binding Domain With Polyphenols: Unveiling Interactions Through Biophysical and In Silico Methods.

The human papillomavirus (HPV) 33 is a high-risk strain that causes lesions with potential cancerous outcomes. Its E2 protein regulates the viral protein transcription and life cycle maintenance. The DNA binding domain (DBD) of the E2 protein plays a crucial role in the viral life cycle. The DBD region of the E2 protein is particularly interesting for targeting and finding potential inhibitors to inhibit its function or dimerization. Given the limited research on HPV 33 and its proteins, the present work delved into the interaction of two natural polyphenolic compounds, resveratrol, and baicalein, with the E2 DBD of HPV 33 using biophysical and in silico studies. Fluorescence studies of the E2 DBD-polyphenol complexes showed fluorescence quenching with a binding constant of the order of 106 M-1. Circular dichroism data reveal conformational changes upon binding with the polyphenols, possibly due to distinct binding sites of the E2 DBD. Differential scanning calorimetry exhibited higher melting temperatures for the two complexes than alone DBD, suggesting the complexes' stability. ITC experiment suggested favorable binding reactions with kd values in the micromolar range. Molecular docking and dynamic simulation studies revealed that the resveratrol binds to the helical region and baicalein near the central dimeric interface of E2 DBD with a good binding affinity, forming a stable protein-ligand complex during the run of 100 ns simulation. Therefore, the current study identifies both polyphenolic compounds as promising candidates for potential antiviral drug development.

SHCBP1 promotes cisplatin resistance of ovarian cancer through AKT/mTOR/Autophagy pathway.

Ovarian cancer caused the highest cancer-related mortality among female reproductive system malignancies. Platinum-based chemotherapy is still the footstone of the chemotherapy for ovarian cancer. However, the molecular mechanisms underlying cisplatin insensitivity and resistance remain unclear. SHC SH2 domain-binding protein 1 (SHCBP1) plays critical roles in the progression and drug resistance of different types of cancer. However, the biological function of SHCBP1 in ovarian cancer progression and cisplatin resistance remains obscure. In this study, we found that SHCBP1 was upregulated in ovarian cancer and the upregulated SHCBP1 has growth-promoting effect on ovarian cancer cells. Furthermore, SHCBP1 silencing sensitize ovarian cancer cells to cisplatin (hereafter referred to as CDDP). Mechanism analysis revealed that SHCBP1 activated the Akt/mTOR pathway and further inhibited autophagy in ovarian cancer cells. Meanwhile, autophagy inhibitors combined with SHCBP1 knockdown enhances CDDP sensitivity. In addition, knockdown of SHCBP1 restricted the proliferation of tumors and increased the cisplatin sensitivity in vivo. These findings suggested that upregulated SHCBP1 promoted the proliferation and CDDP resistance of ovarian cancer. The combination of SHCBP1 inhibition and cisplatin treatment might lead to substantial progress in ovarian cancer targeted therapy.

Myosin-5a facilitates stress granule formation by interacting with G3BP1

Stress granules (SGs) are non-membranous organelles composed of mRNA and proteins that assemble in the cytosol when the cell is under stress. Although the composition of mammalian SGs is both cell-type and stress-dependent, they consistently contain core components, such as Ras GTPase activating protein SH3 domain binding protein 1 (G3BP1). Upon stress, living cells rapidly assemble micrometric SGs, sometimes within a few minutes, suggesting that SG components may be actively transported by the microtubule and/or actin cytoskeleton. Indeed, SG assembly has been shown to depend on the microtubule cytoskeleton and the associated motor proteins. However, the role of the actin cytoskeleton and associated myosin motor proteins remains controversial. Here, we identified G3BP1 as a novel binding protein of unconventional myosin-5a (Myo5a). G3BP1 uses its C-terminal RNA-binding domain to interact with the middle portion of Myo5a tail domain (Myo5a-MTD). Suppressing Myo5a function in mammalian cells, either by overexpressing Myo5a-MTD, eliminating Myo5a gene expression, or treatment with myosin-5 inhibitor, inhibits the arsenite-induced formation of both small and large SGs. This is different from the effect of microtubule disruption, which abolishes the formation of large SGs but enhances the formation of small SGs under stress conditions. We therefore propose that, under stress conditions, Myo5a facilitates the formation of SGs at an earlier stage than the microtubule-dependent process.

A multispecific antibody against SARS-CoV-2 prevents immune escape in vitro and confers prophylactic protection in vivo.

Despite effective countermeasures, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) persists worldwide because of its ability to diversify and evade human immunity. This evasion stems from amino acid substitutions, particularly in the receptor binding domain (RBD) of the spike protein that confers resistance to vaccine-induced antibodies and antibody therapeutics. To constrain viral escape through resistance mutations, we combined antibody variable regions that recognize different RBD sites into multispecific antibodies. Here, we describe multispecific antibodies, including a trivalent trispecific antibody that potently neutralized diverse SARS-CoV-2 variants and prevented virus escape more effectively than single antibodies or mixtures of the parental antibodies. Despite being generated before the appearance of Omicron, this trispecific antibody neutralized all major Omicron variants through BA.4/BA.5 at nanomolar concentrations. Negative stain electron microscopy suggested that synergistic neutralization was achieved by engaging different epitopes in specific orientations that facilitated binding across more than one spike protein. Moreover, a tetravalent trispecific antibody containing the same variable regions as the trivalent trispecific antibody also protected Syrian hamsters against Omicron variants BA.1, BA.2, and BA.5 challenge, each of which uses different amino acid substitutions to mediate escape from therapeutic antibodies. These results demonstrated that multispecific antibodies have the potential to provide broad SARS-CoV-2 coverage, decrease the likelihood of escape, simplify treatment, and provide a strategy for antibody therapies that could help eliminate pandemic spread for this and other pathogens.

Enhancer looping protein LDB1 modulates MYB expression in T-ALL cell lines in vitro by cooperating with master transcription factors

BackgroundDespite significant progress in the prognosis of pediatric T-cell acute lymphoblastic leukemia (T-ALL) in recent decades, a notable portion of children still confronts challenges such as treatment resistance and recurrence, leading to limited options and a poor prognosis. LIM domain-binding protein 1 (LDB1) has been confirmed to exert a crucial role in various physiological and pathological processes. In our research, we aim to elucidate the underlying function and mechanisms of LDB1 within the background of T-ALL.MethodsEmploying short hairpin RNA (shRNA) techniques, we delineated the functional impact of LDB1 in T-ALL cell lines. Through the application of RNA-Seq, CUT&Tag, and immunoprecipitation assays, we scrutinized master transcription factors cooperating with LDB1 and identified downstream targets under LDB1 regulation.ResultsLDB1 emerges as a critical transcription factor co-activator in cell lines derived from T-ALL. It primarily collaborates with master transcription factors (ERG, ETV6, IRF1) to cooperatively regulate the transcription of downstream target genes. Both in vitro and in vivo experiments affirm the essential fuction of LDB1 in the proliferation and survival of cell lines derived from T-ALL, with MYB identified as a significant downstream target of LDB1.ConclusionsTo sum up, our research establishes the pivotal fuction of LDB1 in the tumorigenesis and progression of T-ALL cell lines. Mechanistic insights reveal that LDB1 cooperates with ERG, ETV6, and IRF1 to modulate the expression of downstream effector genes. Furthermore, LDB1 controls MYB through remote enhancer modulation, providing valuable mechanistic insights into its involvement in the progression of T-ALL.

Pharmacological targeting of adaptor proteins in chronic inflammation.

Inflammation, a biological response of the immune system, can be triggered by various factors such as pathogens, damaged cells, and toxic compounds. These factors can lead to chronic inflammatory responses, potentially causing tissue damage or disease. Both infectious and non-infectious agents, as well as cell damage, activate inflammatory cells and trigger common inflammatory signalling pathways, including NF-κB, MAPK, and JAK-STAT pathways. These pathways are activated through adaptor proteins, which possess distinct protein binding domains that connect corresponding interacting molecules to facilitate downstream signalling. Adaptor molecules have gained widespread attention in recent years due to their key role in chronic inflammatory diseases. In this review, we explore potential pharmacological agents that can be used to target adaptor molecules in chronic inflammatory responses. A comprehensive analysis of published studies was performed to obtain information on pharmacological agents. This review highlights thetherapeutic strategies involving small molecule inhibitors, antisense oligonucleotide therapy, and traditional medicinal compounds thathave been found to inhibit the inflammatory response and pro-inflammatory cytokine production. These strategies primarily block the protein-protein interactions in the inflammatory signaling cascade.Nevertheless, extensive preclinical studies and risk assessment methodologies are necessary to ensure their safety.

Monitoring humoral responses against three SARS-CoV-2 vaccines in a university population from Chihuahua, Mexico.

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), the causative agent of coronavirus disease 2019 (COVID-19), has spread worldwide since 2019. Survey of the antibodies against SARS-CoV-2 is one of the most important measures of immunity since it can give an idea on the effectiveness of administered vaccines and the serologic status of individuals. We determined the concentrations of blood IgM and IgG against three SARS-CoV-2 proteins in vaccinated teachers and students among a university population from Chihuahua, Mexico. Humoral response surveillance against the 3C-like proteinase (3CLpro), nuclear protein (NP), and receptor binding domain (RBD) of SARS-CoV-2 was carried out. A total of 239 samples were analyzed: 67 from teachers who were vaccinated with CanSino and 172 from students (27.9% were vaccinated with AstraZeneca, 32.6% with Sinovac, 24.4% with Pfizer-BioNTech, 15.1% with other vaccines). Significant differences in the levels of IgG were observed between serum from individuals prior to vaccination (preimmunization serum) and from those that were vaccinated with CanSino. However, samples from asymptomatic individuals did not show differences between the preimmunization and post-immunization serum. The three vaccinated groups (AstraZeneca, Pfizer and Sinovac) did not show significant differences in anti-RBD IgG antibody titers compared to the positive control group, except for a Pfizer non-COVID-19 subgroup where the level of antibodies in the Pfizer group was 1.7 times higher. Neither vaccine group showed significant differences between those individuals who previously had COVID-19 and uninfected individuals. These results provide a picture of the situation at the time when in-person classes resumed.

Enhanced performance of impedimetric immunosensors to detect SARS-CoV-2 with bare gold nanoparticles and graphene acetic acid

Immunosensors based on electrical impedance spectroscopy allow for label-free, real-time detection of biologically relevant molecules and pathogens, without requiring electro-active materials. Here, we investigate the influence of bare gold nanoparticles (AuNPs), synthesized via laser ablation in solution, on the performance of an impedimetric immunosensor for detecting severe acute respiratory syndrome coronavirus (SARS-CoV-2). Graphene acetic acid (GAA) was used in the active layer for immobilizing anti-SARS-CoV-2 antibodies, owing to its high density of carboxylic groups. Immunosensors incorporating AuNPs exhibited superior performance compared to those relying solely on GAA, achieving a limit of detection (LoD) of 3 x 10−20 g/mL to detect the Spike Receptor Binding Domain (RBD) protein of SARS-CoV-2 and of 2 PFU/mL for inactivated virus. Moreover, these immunosensors presented high selectivity against the H1N1 influenza virus. We anticipate that this platform will be versatile and applicable in the early diagnosis of various diseases and viral infections, thereby facilitating Point-of-Care testing.

Exploring the Therapeutic Potential of Coumarin-thiazolotriazole Pharmacophores for SARS-CoV-2 Spike Protein through In-vitro and In-silico Evaluation.

The pandemic caused by SARS-CoV-2 significantly impacted human life around the globe. Numerous unexpected modifications of the SARS-CoV-2 genome have resulted in the emergence of new types and have caused great concern globally. Inhibitory effects of bioactive phytochemicals derived from natural and synthetic sources are promising for pathogenic viruses. in vitro and in silico techniques were used in the current study to identify novel inhibitors of coumarin clubbed thiazolo[3,2-b][1,2,4]triazoles against the SARS-CoV-2 spike protein. Interestingly, all the tested molecules demonstrated substantial inhibition of spike protein with 91.81-57.90% inhibition. The spike protein was remarkably inhibited by compounds 6k (91.83%), 6j (89.75%), 6m (87.69%),6i (86.60%), 6l (85.40%), 6h (84.70%), 6l (84.70%), 6g (83.40%), 6b (82.60%), 6f (81.90%), while compounds 6d 6a, 6c, and 6e exhibited significant activity against spike protein with 79.60%, 77.10%, 75.30%, and 57.90% inhibition, respectively. The binding mechanism of these novel inhibitors with spike protein was deduced in silico, which reflects that the active molecules firmly bind with the receptor binding domain (RBD) of spike protein, thereby inhibiting its function. The combined in vitro and in silico investigations unfold the therapeutic potential of coumarin-thiazolotriazole scaffolds in the treatment of SARS-CoV-2 infection.

Emerging severe acute respiratory syndrome coronavirus 2 variants and their impact on immune evasion and vaccine-induced immunity.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants harboring mutations in the structural protein, especially in the receptor binding domain (RBD) of spike protein, have raised concern about potential immune escape. The spike protein of SARS-CoV-2 plays a vital role in infection and is an important target for neutralizing antibodies. The mutations that occur in the structural proteins, especially in the spike protein, lead to changes in the virus attributes of transmissibility, an increase in disease severity, a notable reduction in neutralizing antibodies generated and thus a decreased response to vaccines and therapy. The observed multiple mutations in the RBD of the spike protein showed immune escape because it increases the affinity of spike protein binding with the ACE-2 receptor of host cells and increases resistance to neutralizing antibodies. Cytotoxic T-cell responses are crucial in controlling SARS-CoV-2 infections from the infected tissues and clearing them from circulation. Cytotoxic Tcells efficiently recognized the infected cells and killed them by releasing soluble mediator's perforin and granzymes. However, the overwhelming response of Tcells and, subsequently, the overproduction of inflammatory mediators during severe infections with SARS-CoV-2 may lead to poor outcomes. This review article summarizes the impact of mutations in the spike protein of SARS-CoV-2, especially mutations of RBD, on immunogenicity, immune escape and vaccine-induced immunity, which could contribute to future studies focusing on vaccine design and immunotherapy.