Anthraquinone-peptide conjugates as inhibitors of DNA transcription factor binding.

Inhibitors of Activator Protein-1 (AP-1)~ a collective term referring to dimeric transcription factors involved in mediating long term responses to signals that regulate growth control and development~ were designed and synthesised in this study. Truncated peptide sequences of 5-7 residues containing the highly conserved tri-amino acid sequence KCR found in the DNA binding domains of the AP-1 family of proteins were attached to mono-substituted anthraquinones through various amino acid type linkers. It was envisaged that these intercalator-linked peptides would have the advantage of high DNA binding with some sequence selectivity. The peptides (l:ARCKA; 2:AKCRA; 3:AKSRA; 4:AKCRNA; 5:AKCRKA; 6:AKCRNKA; 7:AKCRKRA; 8:AAKCRAA) were synthesised using solid phase peptide synthesis. The amino acid type linkers were attached to 1- chloroanthraquinone Via a nucleophilic displacement reaction and subsequently coupled to the N-terminus of the resin bound peptides using PyBOP. Characterisation of the peptides and the intercalator-linked peptides was conducted using HPLC~ MALDI-MS and 2D-NMR spectroscopy. Biophysical analysis of the peptide conjugates with DNA using fluorimetry and thermal denaturation studies indicated that the AP-I inhibitors were weak intercalators of DNA and suggested that DNA binding of these intercalator-linked peptides was govemed~ in principle, by the peptidic moiety. Electrophoretic mobility shift assays of the peptides and their respective conjugates demonstrated that the latter were more effective in displacing the AP-1 protein compared to the peptide moieties alone. These assays also showed that the peptides and conjugates bearing the sequences AKCRNA (peptide 4) and AKCRKRA (peptide 7) were particularly effective in inhibiting the AP-1 protein binding. Peptides 4 and 7 were also shown to possess an intrinsic propensity to form helical structures upon transfer to hydrophobic environments and also to interact with the DNA AP-I consensus sequence more favourably compared to peptides 1. 2. 3. 5, 6 and 8 in the circular dichroism and molecular modelling studies respectively.