Curdlan sulfate (CRDS), a sulfated 1→3-β-D glucan, previously shown to be a potent HIV entry inhibitor, is characterized in this study as a potent inhibitor of the Dengue virus (DENV). CRDS was identified by in silico blind docking studies to exhibit binding potential to the envelope (E) protein of the DENV. CRDS was shown to inhibit the DENV replication very efficiently in different cells in vitro. Minimal effective concentration of CRDS was as low as 0.1 µg/mL in LLC-MK2 cells, and toxicity was observed only at concentrations over 10 mg/mL. CRDS can also inhibit DENV-1, 3, and 4 efficiently. CRDS did not inhibit the replication of DENV subgenomic replicon. Time of addition experiments demonstrated that the compound not only inhibited viral infection at the host cell binding step, but also at an early post-attachment step of entry (membrane fusion). The direct binding of CRDS to DENV was suggested by an evident reduction in the viral titers after interaction of the virus with CRDS following an ultrafiltration device separation, as well as after virus adsorption to an alkyl CRDS-coated membrane filter. The electron microscopic features also showed that CRDS interacted directly with the viral envelope, and caused changes to the viral surface. CRDS also potently inhibited DENV infection in DC-SIGN expressing cells as well as the antibody-dependent enhancement of DENV-2 infection. Based on these data, a probable binding model of CRDS to DENV E protein was constructed by a flexible receptor and ligand docking study. The binding site of CRDS was predicted to be at the interface between domains II and III of E protein dimer, which is unique to this compound, and is apparently different from the β-OG binding site. Since CRDS has already been tested in humans without serious side effects, its clinical application can be considered.
Background:The applications of transgenic plants in the healthcare system are immense. They offer an alternative strategy for the fabrication of antigenic determinants of medically important pathogens. Objectives: Cloning and transient expression of the vacuolating cytotoxin A (vacA) gene of Helicobacter pylori in Nicotiana benthamiana is undertaken in the present study. Methods: The vacA gene of H. pylori was amplified. The vacA and pBI121 vectors were digested with BamHI and SacI and the vacA gene was cloned in pBI121 by T4 ligation. The vacA-pBI121 construct was transformed into Escherichia coli DH5α and the transformants were confirmed by isolation and sequencing of vacA-pBI121. Further, the vacA-pBI121 was transformed into Agrobacterium tumefaciens EHA105 by electroporation. The transformants were used for agroinfection of N. benthamiana by agroinjection technique and the transgenic plant was screened for vacA gene expression by Sodium Dodecyl Sulphate-Polyacrylamide Gel Electrophoresis (SDS-PAGE). Results: The vacA gene amplification was confirmed by observing an intense DNA band in agarose electrophoresis. Sequencing of vacA gene of E. coli DH5α transformants indicated a gene size of about 2877bp which revealed 99.82% sequence similarity with online available H. pylori vacA gene sequence. The A. tumefaciens EHA105 transformants were confirmed by amplification of the vacA gene. The screening of transgenic leaves of N. benthamiana for vacA gene expression by SDS-PAGE showed VacA protein with a molecular weight of 105kDa. Conclusion: A novel transgenic plant expressing VacA protein was developed as a source of eco-friendly-based synthesis of antigenic determinants for various medical applications.
Leptospirosis is a bacterial zoonotic disease caused by spirochetes of the genus Leptospira that affects human and a wide range of animals. The direct method of diagnosis of leptospirosis, has been so far by culture isolation but it is time consuming and potentially biohazardous. Another traditional method is the detection of antibodies (Serological tests) which is also a time consuming method and fails to identify the infecting serovar. To overcome these limitations associated with the cultivation and serology, we developed PCR assay targeting partial lipL21 gene and lipL41 gene of Leptospires using in-house designed P28/29 and P30/31 primers, with a product size of 385bp and 427bp. The amplicons were subjected to restriction enzyme digestion using RsaI, Pvu II and HindIII for product of P28/29 and ClaI, TaqI and RsaI were used for product of P30/31. The protocols were standardized and the assay targeting the partial lipL21 and lipL41 gene was found to be specific for eight pathogenic Leptospires out of nine leptospires tested. The products were then cloned in pGEMT Easy vector and sequenced to facilitate further studies. PCR could detect the target bacterial gene without any ambiguity and showed good efficiency in detection of targeted species in the sample. This simple, rapid and cost-effective method can be applicable in a prediction system to prevent disease outbreak by these Leptospira species and can be considered as an effective tool for disease diagnosis of Leptospira species.
Thiazoles are notable five-membered heterocyclic rings and their moieties can be found in several biologically active compounds of natural origin, as well as synthetic molecules that possess a wide range of pharmacological activities. Inflammation is the common cause that is associated with different disorders and diseases such as psoriasis, arthritis, infections, asthma, cancer, etc. In this article, the synthesis pattern of these novel molecules are discussed and their anti-inflammatory activities against cyclooxygenase-1 (COX-1), cyclooxygenase-2 (COX-2) and lipoxygenase (LOX) were reviewed and documented. The potent 26 thiazole analogs were validated with molecular docking against main protease (6LU7) and spike binding domain ACE2 receptor (6M0J) to defeat from the COVID-19 infections. Among this, THI-9a showed excellent binding energy and affinity against deadly SAR CoV-2. The reviewed and theoretical study information strongly suggested that thiazole derivatives can be used for the development of futuristic target drugs against death-causing diseases like SAR-CoV-2.
The current pandemic Covid-19 brought about by a newly emerged and highly infectious virus named as Sars-CoV-2 as a worldwide danger, has infected more than 600 million people and number of deaths are continuously rising day by day. Till date there are no medications accessible for treatment. All over the world scientists and researchers are involved in the study of this emerged virus and its lifecycle. Structures of proteins in the life cycle of virus has been revealed in RCSB PDB (Research Collaboratory for Structural Bioinformatics Protein Data Bank) by researchers. Citrus fruits are used to treat many distresses of humans. Literature survey shows that it has various activities. Our research work is meant to identify the phytoconstituents which are having phenolic composition and good antiviral and antioxidant properties from citrus fruits against Covid-19 proteins (spike binding domain with ACE2 receptor and spike binding domain with Main protease) and to know its in-silico molecular basis. In this study, about 25 compounds from citrus fruits which is having a good antiviral and antioxidant properties and also phenolic composition were employed for molecular docking analysis, molecular dynamic simulation studies and ADME studies. Based on present study 2 compounds from Citrus fruits acted well against the Covid-19 proteins. The MD simulations were employed to identify Hesperidin and Procyanidin B2 as hit compounds. Further ADME analysis were studied for top 2 compounds, these compounds can be further taken for in-vitro studies to know the effective activity against Covid-19. HIGHLIGHTS Citrus fruit phytochemicals are fascinating core, which shows an excellent anti-viral, anti-inflammatory and antioxidant properties The SARS-CoV-2 is newly emerged and highly infectious virus, has infected more than 600 million people and number of deaths are continuously rising day by day Molecular docking analysis, Molecular dynamics simulation ADME studies was carried out Compound Hesperidin and Procyanidin B2 showed a good inhibitory activity against SARS-CoV-2 proteins GRAPHICAL ABSTRACT
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