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Drug–DNA interactions: Spectroscopic and footprinting studies of site and sequence specificity of elliptinium
Author(s) -
Mauffret Oliver,
Rene Brigitte,
Convert Odile,
Monnot Monique,
Lescot Elie,
Fermandjian Serge
Publication year - 1991
Publication title -
biopolymers
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.556
H-Index - 125
eISSN - 1097-0282
pISSN - 0006-3525
DOI - 10.1002/bip.360311110
Subject(s) - chemistry , footprinting , intercalation (chemistry) , dna , stereochemistry , base pair , oligonucleotide , dna footprinting , binding site , crystallography , biochemistry , inorganic chemistry , dna binding protein , transcription factor , gene , base sequence
The binding of the antitumoral ellipticine derivative 2‐methyl‐9‐hydroxyellipticinium acetate (elliptinium; NMHE) to DNA was analyzed by the combined use of DNase I foot‐printing and spectroscopic methods. Using two fragments of pBR322 DNA, five discrete NMHE binding sites of 5–7 protected base pairs (bp) were detected by footprinting at 4°C on the analyzed regions. These corresponded to alternating pyrimidines and purines. The inactive derivative 2‐methyl ellipticinium acetate L(NME) lacking a hydroxy group failed to demonstrate DNA protection even at low temperature. Ultraviolet‐absorption and 1 H‐nmr analysis was performed using two autocomplementary octanucleotides d(TGACGTCA) (I) and d(ACTGCAGT) (II). The uv‐absorption titrations resulted in an intercalative binding mode for NMHE in the oligomers. Analysis of the derived biphasic Scatchard plots yielded two binding sites corresponding to ∼6‐bp and 2‐bp sizes and characterized by apparent association constants K 1 ∼ 10 8 M −1 and K 2 ∼ 10 6 M −1 , respectively. The 1 H‐nmr analysis of exchangeable (imino) protons and nonexchangeable protons performed in the one‐ and two‐dimensional modes confirmed the intercalation of NMHE, and further revealed the existence of multiple sites on DNA. Assuming that imino resonance line width concerned the sole kinetic effects, 10‐ms order lifetimes were estimated for the drug–oligonucleotide complexes at 7°C, pH 7, and 0.1 ionic strength. Finally examination of every drug–DNA spectra in the light of the footprinting results indicated that there was a preference for binding of NMHE to the CpG (octamer I) and TpG (octamers I and II) steps.