RESUMEN
Holliday 4-way junctions are key to important biological DNA processes (insertion, recombination, and repair) and are dynamic structures that adopt either open or closed conformations, the open conformation being the biologically active form. Tetracationic metallo-supramolecular pillarplexes display aryl faces about a cylindrical core, an ideal structure to interact with open DNA junction cavities. Combining experimental studies and MD simulations, we show that an Au pillarplex can bind DNA 4-way (Holliday) junctions in their open form, a binding mode not accessed by synthetic agents before. Pillarplexes can bind 3-way junctions too, but their large size leads them to open up and expand that junction, disrupting the base pairing, which manifests in an increased hydrodynamic size and lower junction thermal stability. At high loading, they rearrange both 4-way and 3-way junctions into Y-shaped forks to increase the available junction-like binding sites. Isostructural Ag pillarplexes show similar DNA junction binding behavior but lower solution stability. This pillarplex binding contrasts with (but complements) that of metallo-supramolecular cylinders, which prefer 3-way junctions and can rearrange 4-way junctions into 3-way junction structures. The pillarplexes' ability to bind open 4-way junctions creates exciting possibilities to modulate and switch such structures in biology, as well as in synthetic nucleic acid nanostructures. In human cells, the pillarplexes do reach the nucleus, with antiproliferative activity at levels similar to those of cisplatin. The findings provide a new roadmap for targeting higher-order junction structures using a metallo-supramolecular approach, as well as expanding the toolbox available to design bioactive junction binders into organometallic chemistry.
Asunto(s)
ADN Cruciforme , Ácidos Nucleicos , Humanos , Conformación de Ácido Nucleico , ADN/química , Sitios de UniónRESUMEN
The untranslated regions (UTRs) of viral genomes contain a variety of conserved yet dynamic structures crucial for viral replication, providing drug targets for the development of broad spectrum anti-virals. We combine in vitro RNA analysis with molecular dynamics simulations to build the first 3D models of the structure and dynamics of key regions of the 5' UTR of the SARS-CoV-2 genome. Furthermore, we determine the binding of metallo-supramolecular helicates (cylinders) to this RNA structure. These nano-size agents are uniquely able to thread through RNA junctions and we identify their binding to a 3-base bulge and the central cross 4-way junction located in stem loop 5. Finally, we show these RNA-binding cylinders suppress SARS-CoV-2 replication, highlighting their potential as novel anti-viral agents.
Asunto(s)
Regiones no Traducidas 5' , Antivirales/farmacología , Sustancias Macromoleculares/farmacología , ARN/metabolismo , SARS-CoV-2/efectos de los fármacos , Replicación Viral/efectos de los fármacos , Animales , Antivirales/química , Antivirales/metabolismo , Chlorocebus aethiops , Complejos de Coordinación/química , Complejos de Coordinación/metabolismo , Complejos de Coordinación/farmacología , Genoma Viral/efectos de los fármacos , Sustancias Macromoleculares/química , Sustancias Macromoleculares/metabolismo , Metales Pesados/química , Simulación de Dinámica Molecular , ARN/genética , SARS-CoV-2/química , Células VeroRESUMEN
The untranslated regions (UTRs) of viral genomes contain a variety of conserved yet dynamic structures crucial for viral replication, providing drug targets for the development of broad spectrum anti-virals. We combine in vitro RNA analysis with molecular dynamics simulations to build the first 3D models of the structure and dynamics of key regions of the 5' UTR of the SARS-CoV-2 genome. Furthermore, we determine the binding of metallo-supramolecular helicates (cylinders) to this RNA structure. These nano-size agents are uniquely able to thread through RNA junctions and we identify their binding to a 3-base bulge and the central cross 4-way junction located in stem loop 5. Finally, we show these RNA-binding cylinders suppress SARS-CoV-2 replication, highlighting their potential as novel anti-viral agents.
RESUMEN
A class of rotaxane is created, not by encapsulating a conventional linear thread, but rather by wrapping a large cucurbit[10]uril macrocycle about a three-dimensional, cylindrical, nanosized, self-assembled supramolecular helicate as the axle. The resulting pseudo-rotaxane is readily converted into a proper interlocked rotaxane by adding branch points to the helicate strands that form the surface of the cylinder (like branches and roots on a tree trunk). The supramolecular cylinder that forms the axle is itself a member of a unique and remarkable class of helicate metallo-drugs that bind Y-shaped DNA junction structures and induce cell death. While pseudo-rotaxanation does not modify the DNA-binding properties, proper, mechanically-interlocked rotaxanation transforms the DNA-binding and biological activity of the cylinder. The ability of the cylinder to de-thread from the rotaxane (and thus to bind DNA junction structures) is controlled by the extent of branching: fully-branched cylinders are locked inside the cucurbit[10]uril macrocycle, while cylinders with incomplete branch points can de-thread from the rotaxane in response to competitor guests. The number of branch points can thus afford kinetic control over the drug de-threading and release.
Asunto(s)
ADN/química , Metales/química , Nanoestructuras/química , Rotaxanos/química , Hidrocarburos Aromáticos con Puentes/química , Complejos de Coordinación/química , Imidazoles/química , LigandosRESUMEN
New gold and lipoic based nanocarriers for the delivery of platinum(ii) and platinum(iv) drugs are developed, which allow enhanced loading of the drug on the surface of the nanocarriers and release in a pH-dependent fashion, with superior release at lower pHs which are associated with many tumours. The conjugate nanoparticles and their conjugates enter cells rapidly (within 3 hours). They tend to cluster in vesicles and are also observed by light and electron microscopies in the cytoplasm, endoplasmic reticulum and nucleus. We further incorporate aminoanthraquinone units that are both fluorophores and DNA intercalators. This results in nanocarriers that after drug release will remain surface decorated with DNA-binders challenging the conventional design of the nanocarrier as an inert component. The outcome is nanocarriers that themselves have distinctive, remarkable and unusual DNA binding properties being able to bind and wrap DNA (despite their anionic charge) and provide enhanced cytotoxic activity beyond that conferred by the platinum agents they release. DNA coiling is usually associated with polycations which can disrupt cell membranes; anionic nanoparticles that can cause novel and dramatic effects on DNA may have fascinating potential for new approaches to in-cell nucleic acid recognition. Our findings have implications for the understanding and interpretation of the biological activities of nanoparticles used to deliver other DNA-binding drugs including clinical drug doxorubicin and its formulations.