The current pandemic situation caused by the Betacoronavirus SARS-CoV-2 (SCoV2) highlights the need for coordinated research to combat COVID-19. A particularly important aspect is the development of medication. In addition to viral proteins, structured RNA elements represent a potent alternative as drug targets. The search for drugs that target RNA requires their high-resolution structural characterization. Using nuclear magnetic resonance (NMR) spectroscopy, a worldwide consortium of NMR researchers aims to characterize potential RNA drug targets of SCoV2. Here, we report the characterization of 15 conserved RNA elements located at the 5′ end, the ribosomal frameshift segment and the 3′-untranslated region (3′-UTR) of the SCoV2 genome, their large-scale production and NMR-based secondary structure determination. The NMR data are corroborated with secondary structure probing by DMS footprinting experiments. The close agreement of NMR secondary structure determination of isolated RNA elements with DMS footprinting and NMR performed on larger RNA regions shows that the secondary structure elements fold independently. The NMR data reported here provide the basis for NMR investigations of RNA function, RNA interactions with viral and host proteins and screening campaigns to identify potential RNA binders for pharmaceutical intervention.

A simple-prepare, single-use and cost-effective, in vitro biosensor for the detection of TAR DNA-binding protein 43 (TDP-43), a biomarker of neuro-degenerative disorders, was designed, manufactured and tested. This study reports the first biosensor application for the detection of TDP-43 using a novel biosensor fabrication methodology. Bioconjugation mechanism was applied by conjugating anti-TDP 43 with N-succinimidyl S-acetylthioacetate (SATA) producing a thiol-linked anti-TDP 43, which was used to directly link with gold electrode surface, minimizing the preparation steps for biosensor fabrication and simplifying the biosensor surface. The effectiveness of this bioconjugation mechanism was evaluated and confirmed by FqRRM12 protein, using nuclear magnetic resonance (NMR). The surface coverage of the electrode was analyzed by Time-of-Flight-Secondary Ion Mass Spectrometry (TOF-SIMS). Differential pulse voltammetry (DPV) was acted as the detection transduction mechanism with the use of [Fe(CN)6]3−/4-redox probe. Human TDP-43 peptide of 0.0005 μg/mL to 2 μg/mL in undiluted human serum was analyzed using this TDP-43 biosensor. Interference study of the TDP-43 biosensor using β-amyloid 42 protein and T-tau protein confirmed the specifity this TDP-43 biosensor. This bioconjugation chemistry based approach for biosensor fabrication circumvents tedious gold surface modification and functionalization while enabling specific detection of TDP-43 in less than 1 hr with a low fabrication cost of a single biosensor less than $3.

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A noninvasive, highly sensitive universal immunosensor platform for protein-based biomarker detection is described in this Article. A neutral charged sensing environment is constructed by an antibody with an oppositely charged amino acid as surface charge neutralizer. By adjusting the pH condition of the testing environment, this neutral charged immunosensor (NCI) directly utilizes the electrostatic charges of the analyte for quantification of circulating protein markers, achieving a wide dynamic range covering through the whole picomole level. Comparing with previous studies on electrostatic charges characterization, this NCI demonstrates its capability to analyze not only the negatively charged biomolecules but also positively charged analytes. We applied this NCI for the detection of HE4 antigen with a detection limit at 2.5 pM and Tau antigen with a detection limit at 0.968 pM, demonstrating the high-sensitivity property of this platform. Furthermore, this NCI possesses a simple fabrication method (less than 2 h) and a short testing turnaround time (less than 30 min), providing an excellent potential for further clinical point-of-care applications.