title: DNA sequence recognition in the minor groove by crosslinked polyamides: The effect of N-terminal head group and linker length on binding affinity and specificity.
abstract: Development of sequence-reading polyamides or "lexitropsins" with comparable DNA-binding affinities to cellular proteins raises the possibility of artificially regulated gene expression. Covalent linkage of polyamide ligands, with either a hairpin motif or crosslinking methylene bridge, has greatly improved binding affinity by ensuring their side-by-side register. Whereas hairpin polyamides have been investigated extensively, the optimized structure of crosslinked polyamides remains to be determined. This study examines a series of thiazole-imidazole-pyrrole (TIP) monomers and crosslinked dimers to evaluate the effects on selectivity and binding affinity of different N-terminal head groups attached to the leading thiazole ring and differing methylene linker lengths. Quantitative footprinting of a DNA sequence, containing potential match and mismatch sites for both maximum overlap and one-residue stagger binding modes, allowed measurement of binding constants at each putative site. Within an N-terminal amino TIP series, C7 and C8-linked compounds bound most strongly to these sites, whereas maximum binding affinity was observed for a C6 linker with a formyl head group. A C5 linker gave weak binding with either head group. A hydrogen or acetyl head group abrogated binding. Binding was confirmed by gel shift analyses. The highest specificity for the maximum overlap site over the one-residue stagger was observed with TIP-C7-amino. Selectivity of the leading thiazole was modulated by the head group, with N-terminal formyl TIP exhibiting up to 3-fold specificity for AGT over TGT, suggesting that N-formyl-thiazole may provide sequence discrimination of adenine over thymine. Moreover, the leading head group and methylene linker length significantly influences the binding characteristics of crosslinked polyamides.
authors.affiliation: Cancer Research Campaign Drug-DNA Interactions Research Group, Department of Oncology, Royal Free and University College Medical School, UCL, 91 Riding House Street, London, W1W 7BS, United Kingdom.
authors.forename: C Caroline
authors.initials: CC
authors.lastname: O'Hare
authors.forename: Darcy
authors.initials: D
authors.lastname: Mack
authors.forename: Manju
authors.initials: M
authors.lastname: Tandon
authors.forename: Sanjay K
authors.initials: SK
authors.lastname: Sharma
authors.forename: J William
authors.initials: JW
authors.lastname: Lown
authors.forename: Mary L
authors.initials: ML
authors.lastname: Kopka
authors.forename: Richard E
authors.initials: RE
authors.lastname: Dickerson
authors.forename: John A
authors.initials: JA
authors.lastname: Hartley
mesh_terms: D001665:Binding Sites; D003432:Cross-Linking Reagents / Q000494:pharmacology; D004247:DNA / Q000737:chemistry*; D004268:DNA-Binding Proteins / Q000737:chemistry*; D003850:Deoxyribonuclease I / Q000737:chemistry / Q000378:metabolism; D006838:Hydrocarbons; D006860:Hydrogen Bonding; D007700:Kinetics; D008697:Methane / Q000031:analogs & derivatives* / Q000737:chemistry; D008956:Models, Chemical; D015233:Models, Statistical; D009690:Nucleic Acid Conformation; D009757:Nylons / Q000737:chemistry; D011485:Protein Binding; D012091:Repetitive Sequences, Nucleic Acid
chemical_list: D003432:Cross-Linking Reagents; D004268:DNA-Binding Proteins; D006838:Hydrocarbons; D009757:Nylons; C030011:carbene; D004247:DNA; D003850:Deoxyribonuclease I; D008697:Methane