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1.
Int J Mol Sci ; 23(3)2022 Jan 18.
Article in English | MEDLINE | ID: covidwho-1625435

ABSTRACT

Spike protein of SARS-CoV-2 contains a single-span transmembrane (TM) domain and plays roles in receptor binding, viral attachment and viral entry to the host cells. The TM domain of spike protein is critical for viral infectivity. Herein, the TM domain of spike protein of SARS-CoV-2 was reconstituted in detergent micelles and subjected to structural analysis using solution NMR spectroscopy. The results demonstrate that the TM domain of the protein forms a helical structure in detergent micelles. An unstructured linker is identified between the TM helix and heptapeptide repeat 2 region. The linker is due to the proline residue at position 1213. Side chains of the three tryptophan residues preceding to and within the TM helix important for the function of S-protein might adopt multiple conformations which may be critical for their function. The side chain of W1212 was shown to be exposed to solvent and the side chains of residues W1214 and W1217 are buried in micelles. Relaxation study shows that the TM helix is rigid in solution while several residues have exchanges. The secondary structure and dynamics of the TM domain in this study provide insights into the function of the TM domain of spike protein.


Subject(s)
Detergents/pharmacology , Spike Glycoprotein, Coronavirus/chemistry , Amino Acid Sequence , COVID-19/virology , Cell Membrane/metabolism , Cross-Linking Reagents/pharmacology , Detergents/chemistry , Humans , Magnetic Resonance Spectroscopy , Micelles , Models, Molecular , Nuclear Magnetic Resonance, Biomolecular , Protein Domains/drug effects , Protein Structure, Secondary/drug effects , SARS-CoV-2/chemistry , SARS-CoV-2/metabolism , Spike Glycoprotein, Coronavirus/drug effects , Spike Glycoprotein, Coronavirus/metabolism
2.
Bioorg Chem ; 119: 105550, 2022 02.
Article in English | MEDLINE | ID: covidwho-1561636

ABSTRACT

Infectious diseases caused by new or unknown bacteria and viruses, such as anthrax, cholera, tuberculosis and even COVID-19, are a major threat to humanity. Thus, the development of new synthetic compounds with efficient antimicrobial activity is a necessity. Herein, rationally designed novel multifunctional cationic alternating copolymers were directly synthesized through a step-growth polymerization reaction using a bivalent electrophilic cross-linker containing disulfide bonds and a diamine heterocyclic ring. To optimize the activity of these alternating copolymers, several different diamines and cross-linkers were explored to find the highest antibacterial effects. The synthesized nanopolymers not only displayed good to excellent antibacterial activity as judged by minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) against Staphylococcus aureus, Enterococcus faecalis, Pseudomonas aeruginosa, and Escherichia coli, but also reduced the number of biofilm cells even at low concentrations, without killing mammalian cells. Furthermore, in vivo experiments using infected burn wounds in mice demonstrated good antibacterial activity and stimulated wound healing, without causing systemic inflammation. These findings suggest that the multifunctional cationic nanopolymers have potential as a novel antibacterial agent for eradication of multidrug resistant bacterial infections.


Subject(s)
Anti-Bacterial Agents/pharmacology , Anti-Inflammatory Agents, Non-Steroidal/pharmacology , Biofilms/drug effects , Cations/pharmacology , Polymers/pharmacology , Wound Healing/drug effects , Amines/chemistry , Animals , Bacteria/drug effects , Bacterial Infections/drug therapy , Bacterial Infections/etiology , Burns/complications , COVID-19 , Cell Survival/drug effects , Cross-Linking Reagents , Drug Resistance, Multiple, Bacterial/drug effects , HEK293 Cells/drug effects , Humans , Mice , Microbial Sensitivity Tests , Polymers/chemistry
4.
Proc Natl Acad Sci U S A ; 118(34)2021 08 24.
Article in English | MEDLINE | ID: covidwho-1349700

ABSTRACT

Atomic structures of several proteins from the coronavirus family are still partial or unavailable. A possible reason for this gap is the instability of these proteins outside of the cellular context, thereby prompting the use of in-cell approaches. In situ cross-linking and mass spectrometry (in situ CLMS) can provide information on the structures of such proteins as they occur in the intact cell. Here, we applied targeted in situ CLMS to structurally probe Nsp1, Nsp2, and nucleocapsid (N) proteins from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and obtained cross-link sets with an average density of one cross-link per 20 residues. We then employed integrative modeling that computationally combined the cross-linking data with domain structures to determine full-length atomic models. For the Nsp2, the cross-links report on a complex topology with long-range interactions. Integrative modeling with structural prediction of individual domains by the AlphaFold2 system allowed us to generate a single consistent all-atom model of the full-length Nsp2. The model reveals three putative metal binding sites and suggests a role for Nsp2 in zinc regulation within the replication-transcription complex. For the N protein, we identified multiple intra- and interdomain cross-links. Our integrative model of the N dimer demonstrates that it can accommodate three single RNA strands simultaneously, both stereochemically and electrostatically. For the Nsp1, cross-links with the 40S ribosome were highly consistent with recent cryogenic electron microscopy structures. These results highlight the importance of cellular context for the structural probing of recalcitrant proteins and demonstrate the effectiveness of targeted in situ CLMS and integrative modeling.


Subject(s)
Models, Molecular , SARS-CoV-2/chemistry , Viral Proteins/chemistry , Cross-Linking Reagents/chemistry , HEK293 Cells , Humans , Mass Spectrometry , Protein Domains
5.
Sci Adv ; 7(32)2021 08.
Article in English | MEDLINE | ID: covidwho-1343936

ABSTRACT

Host antibody responses are pivotal for providing protection against infectious agents. We have pioneered a new class of self-assembling micelles based on pentablock copolymers that enhance antibody responses while providing a low inflammatory environment compared to traditional adjuvants. This type of "just-right" immune response is critical in the rational design of vaccines for older adults. Here, we report on the mechanism of enhancement of antibody responses by pentablock copolymer micelles, which act as scaffolds for antigen presentation to B cells and cross-link B cell receptors, unlike other micelle-forming synthetic block copolymers. We exploited this unique mechanism and developed these scaffolds as a platform technology to produce antibodies in vitro. We show that this novel approach can be used to generate laboratory-scale quantities of therapeutic antibodies against multiple antigens, including those associated with SARS-CoV-2 and Yersinia pestis, further expanding the value of these nanomaterials to rapidly develop countermeasures against infectious diseases.


Subject(s)
Antibody Formation , Antigen Presentation/immunology , Cross-Linking Reagents/chemistry , Receptors, Antigen, B-Cell/chemistry , Recombinant Fusion Proteins/immunology , Spike Glycoprotein, Coronavirus/immunology , Yersinia pestis/immunology , Adjuvants, Immunologic , Animals , Female , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Polymers/chemistry , Receptors, Antigen, B-Cell/metabolism
6.
Chem Commun (Camb) ; 57(56): 6871-6874, 2021 Jul 13.
Article in English | MEDLINE | ID: covidwho-1281748

ABSTRACT

The trans-cleavage activity of the target-activated CRISPR/Cas12a liberated an RNA crosslinker from a molecular transducer, which facilitated the assembly of gold nanoparticles. Integration of the molecular transducer with isothermal amplification and CRISPR/Cas12a resulted in visual detection of the N gene and E gene of SARS-CoV-2 in 45 min.


Subject(s)
COVID-19/diagnosis , CRISPR-Cas Systems , Genes, Viral/genetics , Gold/chemistry , Metal Nanoparticles/chemistry , Molecular Diagnostic Techniques/methods , Nucleic Acid Amplification Techniques/methods , SARS-CoV-2/genetics , COVID-19/virology , Colorimetry , Cross-Linking Reagents , RNA/chemistry
7.
Eur J Ophthalmol ; 31(6): 3490-3493, 2021 Nov.
Article in English | MEDLINE | ID: covidwho-1133565

ABSTRACT

PURPOSE: Royal College of Ophthalmologist recent guidance recommended delaying cross-linking services during the COVID-19 pandemic. This study investigates the effects of such delays in the delivery of cross-linking services in patients with keratoconus progression. METHODS: Retrospective observational study of 46 patients with keratoconus progression, whose cross-linking was delayed due to the COVID-19 pandemic. Demographic and clinical details were obtained from assessments on the day of listing, and subsequent review on the day of the procedure. Topographic indices included keratometry of the posterior and anterior corneal surface, maximum keratometry (Kmax), thinnest corneal thickness, ABCD progression and progression based on standard criteria recommendations (1.5 D Kmax & 20 microns thinning). RESULTS: A total of 46 eyes were analysed with an average time between being listed for CXL and having the procedure done was 182 ± 65 days. The delay due to COVID-19 was of 3 months. In this time period they had a significant worsening of all keratometric indices and lost almost one line of visual acuity (0.19 ± 0.19 to 0.26 ± 0.18 LogMAR, p: 0.03). Thirty two eyes (70%) demonstrated progression in accordance with the ABCD progression criteria, while 18 eyes (39%) showed either an increase in Kmax of more than 1.5D or a thinning in corneal thickness of at least 20 µm. CONCLUSIONS: The treatment delay for the keratoconus patients caused further progression and vision worsening. We recommend that corneal collagen crosslinking needs to be considered as a high priority intervention.


Subject(s)
COVID-19 , Keratoconus , Photochemotherapy , Collagen/therapeutic use , Corneal Topography , Cross-Linking Reagents/therapeutic use , Humans , Keratoconus/drug therapy , Pandemics , Photosensitizing Agents/therapeutic use , Riboflavin/therapeutic use , SARS-CoV-2 , Ultraviolet Rays
8.
PLoS Biol ; 18(9): e3000827, 2020 09.
Article in English | MEDLINE | ID: covidwho-807960

ABSTRACT

Matrix proteins are encoded by many enveloped viruses, including influenza viruses, herpes viruses, and coronaviruses. Underneath the viral envelope of influenza virus, matrix protein 1 (M1) forms an oligomeric layer critical for particle stability and pH-dependent RNA genome release. However, high-resolution structures of full-length monomeric M1 and the matrix layer have not been available, impeding antiviral targeting and understanding of the pH-dependent transitions involved in cell entry. Here, purification and extensive mutagenesis revealed protein-protein interfaces required for the formation of multilayered helical M1 oligomers similar to those observed in virions exposed to the low pH of cell entry. However, single-layered helical oligomers with biochemical and ultrastructural similarity to those found in infectious virions before cell entry were observed upon mutation of a single amino acid. The highly ordered structure of the single-layered oligomers and their likeness to the matrix layer of intact virions prompted structural analysis by cryo-electron microscopy (cryo-EM). The resulting 3.4-Å-resolution structure revealed the molecular details of M1 folding and its organization within the single-shelled matrix. The solution of the full-length M1 structure, the identification of critical assembly interfaces, and the development of M1 assembly assays with purified proteins are crucial advances for antiviral targeting of influenza viruses.


Subject(s)
Imaging, Three-Dimensional , Viral Matrix Proteins/chemistry , Amino Acid Sequence , Cross-Linking Reagents/chemistry , Hydrogen-Ion Concentration , Models, Molecular , Mutation/genetics , Protein Multimerization , Protein Structure, Secondary , Protein Subunits/chemistry , Recombination, Genetic/genetics , Viral Matrix Proteins/genetics , Virion/ultrastructure
9.
Med Hypotheses ; 144: 110253, 2020 Nov.
Article in English | MEDLINE | ID: covidwho-753090

ABSTRACT

Coronavirus disease (COVID-19) is a recently discovered infectious disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Graphene is an emerging material due to its extraordinary performance in the field of electronics and antimicrobial textiles. Special attention devoted to graphene oxide-based materials due to its surface to volume ratio is very high which make it easy to attach biomolecules by simple adsorption or by crosslinking between reactive groups and the graphene surface. In response to the COVID-19 pandemic, we have summarized the recent developments of graphene and its derivatives with possible virus detection and textile applications. Moreover, graphene strain sensors can be executed on high-performance textiles and high-throughput drug efficacy screening.


Subject(s)
Biosensing Techniques , COVID-19/prevention & control , Graphite/chemistry , Textiles , Adsorption , Anti-Infective Agents , Cross-Linking Reagents , Electronics , Humans , Materials Testing , Models, Theoretical , Occupational Exposure , Occupational Health , SARS-CoV-2 , Surface Properties , Wearable Electronic Devices
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