Nucleic Acids Res

Site-directed spin labeling is emerging as an essential tool to investigate the structural and dynamical features of RNA. We propose here an enzymatic method, which allows the insertion of a paramagnetic center at a specific position in an RNA molecule. The technique is based on a segmental approach using a ligation protocol with T4 RNA ligase 2. One transcribed acceptor RNA is ligated to a donor RNA in which a thio-modified nucleotide is introduced at its 5'-end by in vitro transcription with T7 RNA polymerase. The paramagnetic thiol-specific reagent is subsequently attached to the RNA ligation product. This novel strategy is demonstrated by introducing a paramagnetic probe into the 55 nucleotides long RNA corresponding to K-turn and Specifier Loop domains from the Bacillus subtilis tyrS T-Box leader RNA. The efficiency of the coupling reaction and the quality of the resulting spin-labeled RNA were assessed by Mass Spectrometry, Electron Paramagnetic Resonance (EPR) and Nuclear Magnetic Resonance (NMR). This method enables various combinations of isotopic segmental labeling and spin labeling schemes, a strategy that will be of particular interest to investigate the structural and dynamical properties of large RNA complexes by NMR and EPR spectroscopies

Potential of EPR spin-trapping to investigate in situ free radicals generation from skin allergens in reconstructed human epidermis: cumene hydroperoxide as proof of concept

The first step in the development of skin sensitisation to a chemical, and in the elicitation offurther allergic contact dermatitis (ACD), is the binding of the allergen to skin proteins after pene-trating into the epidermis. The so-formed antigenic adduct is then recognised by the immunesystem as foreign to the body. Sensitising organic hydroperoxides derived from autoxidation ofnatural terpenes are believed to form antigens through radical-mediated mechanisms, althoughthis has not yet been established. So far,in vitroinvestigations on reactive radical intermediatesderived from these skin sensitisers have been conducted in solution, yet with experimental condi-tions being far away from real-life sensitisation. Herein, we report for the first time, the potentialuse of EPR spin-trapping to study thein situgeneration of free radicals derived from cumenehydroperoxide CumOOH in a 3D reconstructed human epidermis (RHE) model, thus much closerto what may happenin vivo. Among the undesirable effects associated with dermal exposure toCumOOH, it is described to cause allergic and irritant dermatitis, being reported as a significantsensitiser. We considered exploiting the usage of spin-trap DEPMPO as an extensive view of allsort of radicals derived from CumOOH were observed all at once in solution. We showed that inthe EpiskinTMRHE model, both by incubating in the assay medium and by topical application,carbon radicals are mainly formed by redox reactions suggesting the key role of CumOOH-derived carbon radicals in the antigen formation process

Magnetic Properties of Gold Nanoparticles: A Room-Temperature Quantum Effect:

Persistent currents: The magnetism of Au nanoparticles might result from persistent currents. Limited portions of a given sample may even support self‐sustained currents, thus exhibiting remnant magnetization and hysteresis. Observing such a quantum effect at room temperature with user‐friendly samples opens unforeseen possibilities