Multifaceted Aspects of HIV-1 Nucleocapsid Inhibition by TAR-Targeting Peptidyl-Anthraquinones Bearing Terminal Aromatic Moieties

2,6-dipeptidyl-anthraquinones are polycyclic planar systems substituted at opposite ring positions by short aminoacyl side chains. Derivatives with positively charged terminal amino acids showed in vitro inhibition of HIV-1 nucleocapsid (NC) protein correlating with threading intercalation through nucleic acid substrates. We found that the variation of the terminal amino acid into an aromatic moiety has profound effects on the NC inhibition of TAR–RNA melting, granting enhanced interaction with the protein. While all compounds showed appreciable NC and TAR binding, they exhibited different strengths driven by the length of the peptidyl side chains and by the stereochemistry of the terminal tyrosine. Unexpectedly, the best inhibitors of NC-induced TAR melting, characterized by the D- configuration of tyrosine, were able to form ternary complexes without competing with TAR–NC recognition sites, as shown by native mass spectrometry experiments. Furthermore, the hydrophobicity of the terminal residue enhances membrane permeation, with positive implications for further studies on these NC–TAR-targeted compounds.