Calix[4]arenes bearing urea units at the meta position(s) of the upper rim of the macrocyclic skeleton were prepared by the reaction of the corresponding amines with aryl isocyanates. As shown by the 1H NMR and UV/vis titration experiments, these systems are capable of effectively complexing selected anions even in a highly competitive environment (such as DMSO-d6). While the monoureido derivatives showed approximately the same complexation ability irrespective of the substitution (para vs. meta isomers), the bisureas at the upper rim demonstrated interesting differences in complexation. The meta,meta and para,para isomers were shown to prefer 2 : 1 complexes (anion : receptor) regardless of the anion tested, while the analogous meta,para isomer formed 1 : 1 complexes with strongly coordinated anions (e.g. H2PO4-) based on synchronous complexation by both ureido groups. This suggests that the regioselective introduction of urea units into the upper rim of calix[4]arene brings with it the possibility of "tuning" the complexation properties depending on the substitution pattern of the functional groups.

• Publication type
• Journal Article MeSH

The COVID-19 pandemic caused by SARS-CoV-2 virus has created a global damage and has exposed the vulnerable side of scientific research towards novel diseases. The intensity of the pandemic is huge, with mortality rates of more than 6 million people worldwide in a span of 2 years. Considering the gravity of the situation, scientists all across the world are continuously attempting to create successful therapeutic solutions to combat the virus. Various vaccination strategies are being devised to ensure effective immunization against SARS-CoV-2 infection. SARS-CoV-2 spreads very rapidly, and the infection rate is remarkably high than other respiratory tract viruses. The viral entry and recognition of the host cell is facilitated by S protein of the virus. N protein along with NSP3 is majorly responsible for viral genome assembly and NSP12 performs polymerase activity for RNA synthesis. In this study, we have designed a multi-epitope, chimeric vaccine considering the two structural (S and N protein) and two non-structural proteins (NSP3 and NSP12) of SARS-CoV-2 virus. The aim is to induce immune response by generating antibodies against these proteins to target the viral entry and viral replication in the host cell. In this study, computational tools were used, and the reliability of the vaccine was verified using molecular docking, molecular dynamics simulation and immune simulation studies in silico. These studies demonstrate that the vaccine designed shows steady interaction with Toll like receptors with good stability and will be effective in inducing a strong and specific immune response in the body.Communicated by Ramaswamy H. Sarma.

• MeSH
• COVID-19 * prevention & control   MeSH
• Epitopes, B-Lymphocyte MeSH
• Humans MeSH
• Pandemics prevention & control   MeSH
• Reproducibility of Results MeSH
• SARS-CoV-2 metabolism   MeSH
• Molecular Docking Simulation MeSH
• COVID-19 Vaccines MeSH
• Viral Vaccines * chemistry   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH

The human prototypical SR protein SRSF1 is an oncoprotein that contains two RRMs and plays a pivotal role in RNA metabolism. We determined the structure of the RRM1 bound to RNA and found that the domain binds preferentially to a CN motif (N is for any nucleotide). Based on this solution structure, we engineered a protein containing a single glutamate to asparagine mutation (E87N), which gains the ability to bind to uridines and thereby activates SMN exon7 inclusion, a strategy that is used to cure spinal muscular atrophy. Finally, we revealed that the flexible inter-RRM linker of SRSF1 allows RRM1 to bind RNA on both sides of RRM2 binding site. Besides revealing an unexpected bimodal mode of interaction of SRSF1 with RNA, which will be of interest to design new therapeutic strategies, this study brings a new perspective on the mode of action of SRSF1 in cells.

• MeSH
• Asparagine genetics   MeSH
• Exons genetics   MeSH
• HEK293 Cells MeSH
• Glutamic Acid genetics   MeSH
• Humans MeSH
• RNA Splice Sites genetics   MeSH
• RNA Recognition Motif genetics   MeSH
• Nuclear Magnetic Resonance, Biomolecular MeSH
• Survival of Motor Neuron 1 Protein genetics   MeSH
• Protein Engineering MeSH
• Recombinant Proteins genetics   isolation & purification   metabolism   ultrastructure   MeSH
• Serine-Arginine Splicing Factors genetics   isolation & purification   metabolism   ultrastructure   MeSH
• RNA Splicing * MeSH
• Molecular Dynamics Simulation MeSH
• Muscular Atrophy, Spinal genetics   therapy   MeSH
• Amino Acid Substitution MeSH
• Uridine metabolism   MeSH
• Computational Biology MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Nanoparticles functionalized with specific biological recognition molecules play a major role for sensor response enhancement in surface plasmon resonance (SPR) based biosensors. The functionalization procedure of such nanoparticles is crucial, since it influences their interactions with the environment and determines their applicability to biomolecular detection in complex matrices. In this work we show how the ζ-potential (Zpot) of bio-functionalized gold spherical NPs (Bio-NPs) is related to the SPR sensor response enhancement of an immune-sandwich-assay for the detection of the carcinoembryonic antigen (CEA), a cancer marker for colorectal carcinomas. In particular, we prepare bio-functional nanoparticles by varying the amount of peptide (either streptavidin or antibody against CEA) bound on their surface. Specific and non-specific sensor responses, reproducibility, and colloidal stability of those bio-functional nanoparticles are measured via SPR and compared to ζ-potential values. Those parameters are first measured in buffer solution, then measured again when the surface of the biosensor is exposed to blood plasma, and finally when the nanoparticles are immersed in blood plasma and flowed overnight on the biosensor. We found that ζ-potential values can guide the design of bio-functional NPs with improved binding efficiency and reduced non-specific sensor response, suitable reproducibility and colloidal stability, even in complex matrixes like blood plasma.

• MeSH
• Hematologic Tests methods   MeSH
• Carcinoembryonic Antigen * blood   MeSH
• Plasma chemistry   MeSH
• Humans MeSH
• Biomarkers, Tumor analysis   MeSH
• Nanoparticles analysis   MeSH
• Peptides analysis   MeSH
• Surface Plasmon Resonance methods   MeSH
• Gold analysis   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Alzheimer's disease (AD) is one of the most significant neurodegenerative disorders and its symptoms mostly appear in aged people. Catechol-o-methyltransferase (COMT) is one of the known target enzymes responsible for AD. With the use of 23 known inhibitors of COMT, a query has been generated and validated by screening against the database of 1500 decoys to obtain the GH score and enrichment value. The crucial features of the known inhibitors were evaluated by the online ZINC Pharmer to identify new leads from a ZINC database. Five hundred hits were retrieved from ZINC Pharmer and by ADMET (absorption, distribution, metabolism, excretion, and toxicity) filtering by using FAF-Drug-3 and 36 molecules were considered for molecular docking. From the COMT inhibitors, opicapone, fenoldopam, and quercetin were selected, while ZINC63625100_413 ZINC39411941_412, ZINC63234426_254, ZINC63637968_451, and ZINC64019452_303 were chosen for the molecular dynamics simulation analysis having high binding affinity and structural recognition. This study identified the potential COMT inhibitors through pharmacophore-based inhibitor screening leading to a more complete understanding of molecular-level interactions.

• MeSH
• Alzheimer Disease drug therapy   enzymology   physiopathology   MeSH
• Gene Expression MeSH
• Databases, Pharmaceutical MeSH
• Catechol O-Methyltransferase Inhibitors chemistry   pharmacology   MeSH
• Protein Interaction Domains and Motifs MeSH
• Catechol O-Methyltransferase chemistry   MeSH
• Kinetics MeSH
• Protein Conformation, alpha-Helical MeSH
• Protein Conformation, beta-Strand MeSH
• Humans MeSH
• Ligands MeSH
• Nootropic Agents chemistry   pharmacology   MeSH
• High-Throughput Screening Assays * MeSH
• Molecular Dynamics Simulation MeSH
• Molecular Docking Simulation MeSH
• Substrate Specificity MeSH
• Protein Structure, Tertiary MeSH
• Thermodynamics MeSH
• Protein Binding MeSH
• Binding Sites MeSH
• Structure-Activity Relationship MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH

The RNA recognition motif (RRM) is the most common RNA binding domain across eukaryotic proteins. It is therefore of great value to engineer its specificity to target RNAs of arbitrary sequence. This was recently achieved for the RRM in Rbfox protein, where four mutations R118D, E147R, N151S, and E152T were designed to target the precursor to the oncogenic miRNA 21. Here, we used a variety of molecular dynamics-based approaches to predict specific interactions at the binding interface. Overall, we have run approximately 50 microseconds of enhanced sampling and plain molecular dynamics simulations on the engineered complex as well as on the wild-type Rbfox·pre-miRNA 20b from which the mutated systems were designed. Comparison with the available NMR data on the wild type molecules (protein, RNA, and their complex) served to establish the accuracy of the calculations. Free energy calculations suggest that further improvements in affinity and selectivity are achieved by the S151T replacement.

• MeSH
• Nucleic Acid Conformation MeSH
• Humans MeSH
• MicroRNAs chemistry   genetics   metabolism   MeSH
• Models, Molecular MeSH
• RNA Recognition Motif * genetics   MeSH
• Nuclear Magnetic Resonance, Biomolecular MeSH
• Protein Engineering MeSH
• RNA-Binding Proteins chemistry   genetics   metabolism   MeSH
• RNA chemistry   metabolism   MeSH
• Amino Acid Sequence MeSH
• Molecular Dynamics Simulation MeSH
• RNA Stability MeSH
• Protein Binding MeSH
• Binding Sites genetics   MeSH
• Computational Biology MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

• MeSH
• Host-Parasite Interactions MeSH
• Publication type
• Abstracts MeSH
• Congress MeSH
• Program MeSH
• Collected Work MeSH
• News MeSH

• Conspectus
• Patologie. Klinická medicína
• NML Fields
• infekční lékařství

2'-Deoxyribonucleoside triphosphates (dNTPs) containing 5-(hydroxymethyl)cytosine (5hmC) protected with photocleavable groups (2-nitrobenzyl or 6-nitropiperonyl) were prepared and studied as substrates for the enzymatic synthesis of oligonucleotides and DNA containing a photocaged epigenetic 5hmC base. DNA probes containing photocaged or free 5hmC in the recognition sequence of restriction endonucleases were prepared and used for the study of the photorelease of caged DNA by UV or visible light at different wavelengths. The nitrobenzyl-protected dNTP was a slightly better substrate for DNA polymerases in primer extension or PCR, whereas the nitropiperonyl-protected nucleotide underwent slightly faster photorelease at 400 nm. However, both photocaged building blocks can be used in polymerase synthesis and the photorelease of 5hmC in DNA.

• MeSH
• 5-Methylcytosine analogs & derivatives   chemical synthesis   chemistry   MeSH
• Deoxyribonucleosides chemical synthesis   chemistry   MeSH
• DNA chemical synthesis   chemistry   MeSH
• Photochemical Processes MeSH
• Polyphosphates chemical synthesis   chemistry   MeSH
• Light MeSH
• Ultraviolet Rays MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Nucleosides, nucleotides and 2'-deoxyribonucleoside triphosphates (dNTPs) containing 5-(hydroxymethyl)uracil protected with photocleavable groups (2-nitrobenzyl-, 6-nitropiperonyl or 9-anthrylmethyl) were prepared and tested as building blocks for the polymerase synthesis of photocaged oligonucleotides and DNA. Photodeprotection (photorelease) reactions were studied in detail on model nucleoside monophosphates and their photoreaction quantum yields were determined. Photocaged dNTPs were then tested and used as substrates for DNA polymerases in primer extension or PCR. DNA probes containing photocaged or free 5-hydroxymethylU in the recognition sequence of restriction endonucleases were prepared and used for the study of photorelease of caged DNA by UV or visible light at different wavelengths. The nitropiperonyl-protected nucleotide was found to be a superior building block because the corresponding dNTP is a good substrate for DNA polymerases, and the protecting group is efficiently cleavable by irradiation by UV or visible light (up to 425 nm).

• MeSH
• DNA-Directed DNA Polymerase metabolism   MeSH
• DNA biosynthesis   chemistry   MeSH
• Photochemical Processes * MeSH
• Nucleic Acid Conformation MeSH
• Models, Molecular MeSH
• Nucleotides chemistry   MeSH
• Pentoxyl analogs & derivatives   chemistry   MeSH
• Light * MeSH
• Publication type
• Journal Article MeSH

The Fox-1 RNA recognition motif (RRM) domain is an important member of the RRM protein family. We report a 1.8 Å X-ray structure of the free Fox-1 containing six distinct monomers. We use this and the nuclear magnetic resonance (NMR) structure of the Fox-1 protein/RNA complex for molecular dynamics (MD) analyses of the structured hydration. The individual monomers of the X-ray structure show diverse hydration patterns, however, MD excellently reproduces the most occupied hydration sites. Simulations of the protein/RNA complex show hydration consistent with the isolated protein complemented by hydration sites specific to the protein/RNA interface. MD predicts intricate hydration sites with water-binding times extending up to hundreds of nanoseconds. We characterize two of them using NMR spectroscopy, RNA binding with switchSENSE and free-energy calculations of mutant proteins. Both hydration sites are experimentally confirmed and their abolishment reduces the binding free-energy. A quantitative agreement between theory and experiment is achieved for the S155A substitution but not for the S122A mutant. The S155 hydration site is evolutionarily conserved within the RRM domains. In conclusion, MD is an effective tool for predicting and interpreting the hydration patterns of protein/RNA complexes. Hydration is not easily detectable in NMR experiments but can affect stability of protein/RNA complexes.

• MeSH
• Crystallography, X-Ray MeSH
• Humans MeSH
• RNA Recognition Motif genetics   MeSH
• Mutagenesis, Site-Directed MeSH
• Nuclear Magnetic Resonance, Biomolecular MeSH
• Recombinant Proteins chemistry   genetics   metabolism   MeSH
• RNA metabolism   MeSH
• RNA Splicing Factors chemistry   genetics   metabolism   MeSH
• Molecular Dynamics Simulation MeSH
• Amino Acid Substitution MeSH
• Binding Sites MeSH
• Water chemistry   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH