Sequence-specific recognition of DNA from the minor groove

Abstract

Footprinting and fluorescence melting techniques were utilised in this thesis to study a range of novel DNA-binding ligands to increase the knowledge of this class of compounds.  So as to assess the binding selectivity of DNA binding ligands, four minimal length novel footprinting sequences were designed, containing every di-, tri- and symmetrical hexa-nucleotide step (the latter split between two fragments).

The novel universal DNA fragments were then combined with ligand-specific fragments to study the binding of novel analogues of the bis-intercalator TANDEM to the dinucleotide TpA, especially when flanked by AT-base pairs; binding was attenuated or abolished by removing the disulphide cross-bridge or replacing the quinoxaline groups with naphthylene rings.  However, the binding affinity was increased by 3040-fold by substituting the two value residues in the octadepsipeptide ring for lysines.

Polyamides are a group of DNA minor groove ligands with established binding rules for selectivity recognising all four base pair combination through the use of pyrrole, imidazole and hydroxypyrrole rings.  The sequence selectivity of a range of polyamide ligands containing novel ring moieties was examined with a view to improving their selectivity and affinity.  It was found that C-terminal pyridoimidazole or benzimidazole ring systems increased the affinity for A or T compared to a pyrrole group.  The position of pyridoimidazoles and benzimidazoles in 2:1 dimers is also important, with pyridoimidazole creating a staggered dimer.  An isopropyl-thiazole ring system was shown to target guanine selectively, while also creating a staggered 2:1 complex.  The introduction of an imidazole to the polyamide in place of a pyrrole decreased the binding affinity significantly, as well as altered the sequence selectivity.

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