Biology subjects

Minari, J. B.

Publications and source records attributed to Minari, J. B..

4 recordsLinked to original sources

bioRxiv2025

Computational Analysis of Silent Mutation Effects on SARS-CoV-2 RNA-Host RNA-Binding Protein Interactome

Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=138 SRC="FIGDIR/small/677528v1_ufig1.gif" ALT="Figure 1"> View larger version (27K): org.highwire.dtl.DTLVardef@131df69org.highwire.dtl.DTLVardef@1429b6corg.highwire.dtl.DTLVardef@1ea0eacorg.highwire.dtl.DTLVardef@1696837_HPS_FORMAT_FIGEXP M_FIG C_FIG AbstractRNA-Binding proteins (RBPs) play critical roles in host-virus interaction. They facilitate the regulation of viral RNA (vRNA) turnover by recognizing and forming complexes with the vRNA structure via specific RNA motifs-RNA binding domain interaction. However, due to consistent evolving nature of viruses, silent mutations in the viral genome can impact RBP-vRNA binding thereby altering the RNA processing. While efforts have been made in characterizing other forms of mutations leading to changes in amino acids sequence in SARS-CoV-2 variants, details on how silent mutations impact RBP-vRNA interaction remain limited. Here, we use extensive in silico mutagenesis to introduce silent mutations in the SARS-CoV-2 genome to generate four different synthetic variants and map the interaction of the variants and the wild-type with a catalogue of human RBPs. Our result shows variation in accumulation and reduction of the RBPs binding motifs in the variants compared to the virus reference sequence on a global scale and at the UTRs. The majority of the RBPs with AU-rich binding motifs are reduced in the variants, while RBPs with mostly GC-rich motifs accumulate more binding positions, suggesting that a single change from U/A to G/C and vice versa can impact RBP- viral interactions. Furthermore, we use structural analysis to show the interaction of the vRNA with PUF60 and KHDRBS3 proteins, two RBPs that have not been previously implicated in SARS-CoV- 2 interactome. Our findings show that loss to the conserved poly(U) in PUF60 binding motifs in some of the variants affects its interaction with the protein at the 5' end, which may disrupt the function of the protein as an anti-viral RNA regulator. We also predicted the key residues in KHDRBS3 interacting with its binding motif in the wild-type at the 3' end, while noting that the vRNA structural changes in the variants may contribute to the loss of this interaction. Overall, our predictions contribute to the insights into virus evolution and pathogenicity of potential new variants due to the impact of synonymous changes in the nucleotide sequences on protein-RNA interaction.

bioinformatics↗

bioRxiv2024

Impact of different synonymous codon substitution strategies on SARS-CoV-2 nucleocapsid protein expression in Escherichia coli

Synonymous codon substitution, a gene engineering approach in synthetic biology, has been effective in improving the codon composition of recombinant genes of interest based on various criteria without altering the amino acid sequence. The SARS-CoV-2 virus nucleocapsid (N) protein is a stable, conserved and highly immunogenic that is less prone to mutation during infection, making it a key antigen in in vitro diagnosis, vaccine development, immunological and structural studies. While reports have focused on applying optimized N protein for different applications, the basic parameters used by different optimization tools for choosing the best approach for the N gene synonymous codon substitution are often neglected. Here, we analyzed the influence of different synonymous codon substitution strategies on SARS-CoV-2 N-protein expression in E. coli. Using different codon optimization (CO) and harmonization (CH) tools, we predicted and compared how parameters such as GC content, Codon Adaptation Index, codon quality and number of rare codons present in these sequences affect the N-protein expression. Our results also show that Minimum Free Energy (MFE) and RNA structure of N-term and C-tail of the N-protein coding sequence influence protein folding. We then predicted that the SR-rich region of the N-protein may contribute to slowing down the elongation rate during translation. This work presents a fundamental analysis of how different optimization tools affect SARS-CoV-2 N-protein expression and folding and suggests a basic approach to choosing the best strategy for optimal expression and folding of the protein for further studies. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=60 SRC="FIGDIR/small/622014v3_ufig1.gif" ALT="Figure 1"> View larger version (15K): org.highwire.dtl.DTLVardef@1fbd7f8org.highwire.dtl.DTLVardef@11fe81borg.highwire.dtl.DTLVardef@1bf787borg.highwire.dtl.DTLVardef@17edc57_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsCodon substitution affects SARS-CoV-2 nucleocapsid (N) protein expression. SARS-CoV-2 N-protein expression varies with different codon optimization tools. RNA structures of N- and C-term impact RNA stability, protein expression and folding. SR-rich region of the N-protein may slow down elongation rate during translation.

bioinformatics↗

bioRxiv2024

Green synthesized silver nanoparticles from Moringa: Potential for preventative treatment of SARS-CoV-2 contaminated water

Biogenic silver nanoparticles have been reported as good antimicrobial candidates. Thus, this study investigated the antiviral activity of silver nanoparticles synthesized against SARS-CoV-2. The silver nanoparticle was biosynthesized using leave extracts of Moringa oleifera (AgNPmo) and characterized using UV-Vis spectroscopy, Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy, and X-ray diffractometry (XRD). The AgNPmo was first tested on clinical bacterial isolates, Pseudomonas aeruginosa (ATTC 154423) and Staphylococcus aureus (ATTC 209233), to ascertain its antimicrobial potential. In-vitro studies were also conducted to determine the cytotoxicity effect of AgNPmo on Vero cells. The efficacy and concentration of AgNPmo against SARS-CoV-2 were evaluated using a qPCR assay in a dose-dependent manner. The results demonstrated the successful biosynthesis and characterization of AgNPmo and its efficacy against the bacterial isolates. The AgNPmo showed low toxicity on the Vero cells. The IC50 from the cytotoxicity assay demonstrated the antiviral activity of the AgNPmo on the SARS-CoV-2 virus, leading to an increase in the Cycle threshold values, notably at 48 hours of incubation and at low concentrations. The results showed that the biogenic AgNPmo synthesized was cost-effective and showed both antimicrobial and antiviral potentials. These findings suggest that the nanoparticles could be a promising alternative for combating SARS-CoV-2, especially for water purification and preventing transmission.

microbiology↗

bioRxiv2024

CRISPR-CAS9 MEDIATED EDITING OF THE LACZ GENE FOR OPTIMIZING LIPASE PRODUCTION AND CHARACTERIZATION IN ENGINEERED ESCHERICHIA COLI USING POTATO PEEL AS A GLUCOSE SOURCE

Improper disposal of potato peels harms the environment, wasting nutrient-rich resources that could be used for beneficial enzyme production like lipase. This study aimed to edit the lacZ gene in Escherichia coli using utilizing the CRISPR Cas9 technology. Both edited and unedited E. coli were used for submerged fermentation of potato peels to produce and characterize lipase. The lacZ gene was edited using the CRISPR Cas9 technology, and the efficiency was measured using multiplex PCR and gel electrophoresis. To measure lipolytic activity, olive oil screening was performed. The temperature and pH of the lipase were used to characterize it after partial purification and submerged fermentation at 10{degrees}C, 30{degrees}C, and 45{degrees}C. Blue colonies indicated the lacZ gene was unedited, lacZ gene editing and repair was demonstrated by white colonies, and no colonies demonstrated the editing but not repair of the lacZ gene. Bands at 1,100 bp indicated unedited lacZ gene, while 650 bp showed edited lacZ gene. Increased cell mass was observed at 10{degrees}C. CRISPR Cas9 edited E. coli showed a clearer zone than the unedited in the lipase medium. The highest lipase activity from both edited and unedited E. coli was at 35{degrees}C, and the lowest at 65{degrees}C. Optimal pH was 7, with lowest activity at pH 4. The CRISPR Cas9 edited E. coli demonstrated significantly higher enzyme activities (p<0.05). This study concluded that CRISPR Cas9-mediated lacZ gene editing in E. coli enhances its ability to utilize potato peels, increasing lipase production.

molecular biology↗