Resolving a guanine-quadruplex structure in the SARS-CoV-2 genome through circular dichroism and multiscale molecular modeling.

The genome of SARS-CoV-2 coronavirus is made up of a single-stranded RNA fragment that can assume a specific secondary structure, whose stability can influence the virus's ability to reproduce. Recent studies have identified putative guanine quadruplex sequences in SARS-CoV-2 genome fragments that are involved in coding for both structural and non-structural proteins. In this contribution, we focus on a specific G-rich sequence referred to as RG-2, which codes for the non-structural protein 10 (Nsp10) and assumes a guanine-quadruplex (G4) arrangement. We provide the secondary structure of RG-2 G4 at atomistic resolution by molecular modeling and simulation, validated by the superposition of experimental and calculated electronic circular dichroism spectra. Through both experimental and simulation approaches, we have demonstrated that pyridostatin (PDS), a widely recognized G4 binder, can bind to and stabilize RG-2 G4 more strongly than RG-1, another G4 forming sequence that was previously proposed as a potential target for antiviral drug candidates. Overall, this study highlights RG-2 as a valuable target to inhibit the translation and replication of SARS-CoV-2, paving the way towards original therapeutic approaches against emerging RNA viruses.

How the Metal Ion Affects the 1H NMR Chemical Shift Values of Schiff Base Metal Complexes: Rationalization by DFT Calculations.

The chemical shift (CS) values obtained by 1H NMR spectroscopy for the hydrogen atoms of a tetradentate N2O2-substituted Salphen ligand (H2L1) are differently shifted in its complexes of nickel(II), palladium(II), platinum(II), and zinc(II), all bearing the same charge on the metal ions. To rationalize the observed trends, DFT calculations have been performed in the implicit d6-DMSO solvent in terms of the electronic effects induced by the metal ion and of the nature and strength of the metal-N and metal-O bonds. Overall, the results obtained point out that, in the complexes involving group 10 elements, the CS values show the greater shift when considering the two hydrogen atoms at a shorter distance from the coordinated metal center and follow the decreasing metal charge in the order Ni > Pd > Pt. This trend suggests a more covalent character of the ligand-metal bonds with the increase of the metal atomic number. Furthermore, a slightly poorer agreement between experimental and calculated data is observed in the presence of the nickel(II) ion. Such discrepancy is explained by the formation of stacked oligomers, aimed at minimizing the repulsive interactions with the polar DMSO solvent.

Mechanism of the Covalent Inhibition of Human Transmembrane Protease Serine 2 as an Original Antiviral Strategy.

The Transmembrane Protease Serine 2 (TMPRSS2) is a human enzyme which is involved in the maturation and post-translation of different proteins. In addition to being overexpressed in cancer cells, TMPRSS2 plays a further fundamental role in favoring viral infections by allowing the fusion of the virus envelope with the cellular membrane, notably in SARS-CoV-2. In this contribution, we resort to multiscale molecular modeling to unravel the structural and dynamical features of TMPRSS2 and its interaction with a model lipid bilayer. Furthermore, we shed light on the mechanism of action of a potential inhibitor (nafamostat), determining the free-energy profile associated with the inhibition reaction and showing the facile poisoning of the enzyme. Our study, while providing the first atomistically resolved mechanism of TMPRSS2 inhibition, is also fundamental in furnishing a solid framework for further rational design targeting transmembrane proteases in a host-directed antiviral strategy.

Predicting the Three-Dimensional Structure of the c-KIT Proto-Oncogene Promoter and the Dynamics of Its Strongly Coupled Guanine Quadruplexes.

Guanine quadruplexes (G4s) play essential protective and regulatory roles within cells, influencing gene expression. In several gene-promoter regions, multiple G4-forming sequences are in close proximity and may form three-dimensional arrangements. We analyze the interplay among the three neighboring G4s in the c-KIT proto-oncogene promoter (WK1, WSP, and WK2). We highlight that the three G4s are structurally linked and their cross-talk favors the formation of a parallel structure for WSP. Relying on all-atom molecular dynamic simulations exceeding the µs time scale and using enhanced sampling methods, we provide the first computationally resolved structure of a well-organized G4 cluster in the promoter of a crucial gene involved in cancer development. Our results indicate that neighboring G4s influence their mutual three-dimensional arrangement and provide a powerful tool to predict and interpret complex DNA structures that can ultimately be used as a starting point for drug discovery.

Understanding the Interactions of Guanine Quadruplexes with Peptides as Novel Strategies for Diagnosis or Tuning Biological Functions.

Guanine quadruplexes (G4s) are nucleic acid structures exhibiting a complex structural behavior and exerting crucial biological functions in both cells and viruses. The specific interactions of peptides with G4s, as well as an understanding of the factors driving the specific recognition are important for the rational design of both therapeutic and diagnostic agents. In this review, we examine the most important studies dealing with the interactions between G4s and peptides, highlighting the strengths and limitations of current analytic approaches. We also show how the combined use of high-level molecular simulation techniques and experimental spectroscopy is the best avenue to design specifically tuned and selective peptides, thus leading to the control of important biological functions.

Salphen metal complexes as potential anticancer agents: interaction profile and selectivity studies toward the three G-quadruplex units in the KIT promoter.

DNA G-rich sequences can organize in four-stranded structures called G-quadruplexes (G4s). These motifs are enriched in significant sites within the human genomes, including telomeres and promoters of cancer related genes. For instance, KIT proto-oncogene promoter, associated with diverse cancers, contains three adjacent G4 units, namely Kit2, SP, and Kit1. Aiming at finding new and selective G-quadruplex binders, we have synthesized and characterized five non-charged metal complexes of Pt(II), Pd(II), Ni(II), Cu(II) and Zn(II) of a chlorine substituted Salphen ligand. The crystal structure of the Pt(II) and Pd(II) complexes was determined by XRPD. FRET measurements indicated that Pt(II) and Pd(II) compounds stabilize Kit1 and Kit2 G4s but not SP, telomeric and double stranded DNA. Spectroscopic investigations (UV-Vis, circular dichroism and fluorescence) suggested the Cu(II) complex as the most G4-selective compound. Interestingly, docking simulations indicate that the synthesized compounds fit groove binding pockets of both Kit1 and Kit2 G4s. Moreover, they exhibited dose-dependent cytotoxic activity in MCF-7, HepG2 and HeLa cancer cells.