Minor-groove recognition of the self-complementary duplex d(CGCGAATTCGCG)2 by Hoechst 33258: a high-field NMR study.
The interaction of Hoechst 33258, a fluorescent DNA stain, has been studied by using the synthetic, self-complementary oligonucleotide duplex d(CGCGAATTCGCG)2. Spectrofluorometric Scatchard analysis indicated that there was only a single class of binding site and that the 1:1 complex had a dissociation constant of (3.47 +/- 0.1) X 10(-6) M at 25 degrees C. Spectroscopic titration by high-field 1H NMR confirmed the 1:1 complex and by means of ID and 2D (NOESY, COSY) techniques the binding site was defined as the minor groove formed by the AATT stretch. Plentiful cross-peaks were measurable and resonance doubling occurred because of the lifting of the diad symmetry of the oligonucleotide on ligand binding. Many individual resonances of both strands of the DNA could be assigned for the complex because of these features, along with the occurrence of slow exchange on the NMR time scale. The results of this NMR spectroscopic solution study were compared with those of previous X-ray crystallographic studies of the same complex. From nuclear Overhauser effect data measured for the complex, a detailed three-dimensional model was constructed with the aid of molecular graphics.
Department of Pharmacy, University of Manchester, U.K.
J A
JA
Parkinson
J
J
Barber
K T
KT
Douglas
J
J
Rosamond
D
D
Sharples
D001483:Base Sequence; D001665:Binding Sites; D006690:Bisbenzimidazole / Q000378:metabolism*; D004247:DNA / Q000737:chemistry; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008968:Molecular Conformation; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism*
D009838:Oligodeoxyribonucleotides; D004247:DNA; D006690:Bisbenzimidazole