Identification of preferred distamycin-DNA binding sites by the combinatorial method REPSA.
The combinatorial method restriction endonuclease protection, selection, and amplification (REPSA) was used to determine the preferred duplex DNA binding sites of the peptide N-methylpyrrolecarboxamide antibiotic distamycin A. After 12 rounds of REPSA, several sequences were identified that bound distamycin with an apparent affinity of 2-20 nM. Among these, the highest-affinity sites averaged 10 bp in length, suggesting that these sites may be occupied by multiple, cooperatively interacting distamycin molecules. Presently, REPSA is the only combinatorial approach that allows the identification of preferred DNA targets for small molecule ligands at physiologically relevant concentrations in solution. As such, it should prove useful in the design and screening of sequence-specific DNA-binding molecules.
Hardenbol
P
P
Department of Tumor Biology, University of Texas M. D. Anderson Cancer Center, Houston 77030, USA.
Wang
J C
JC
Van Dyke
M W
MW
D000900:Anti-Bacterial Agents / Q000737:chemistry*; D001483:Base Sequence; D001665:Binding Sites; D004247:DNA / Q000737:chemistry*; D018983:DNA Footprinting; D004262:DNA Restriction Enzymes; D004214:Distamycins / Q000378:metabolism*; D008024:Ligands; D008969:Molecular Sequence Data; D021141:Nucleic Acid Amplification Techniques; D013698:Templates, Genetic
D000900:Anti-Bacterial Agents; D004214:Distamycins; D008024:Ligands; D004247:DNA; D004262:DNA Restriction Enzymes