Nucleic Acids Research, 2002, Vol. 30, No. 8 1774-1781
© 2002 Oxford University Press
DNA sequence recognition by the indolocarbazole antitumor antibiotic AT2433-B1 and its diastereoisomer
Laboratoire de Pharmacologie Antitumorale du Centre Oscar Lambret and INSERM U-524, IRCL, Place de Verdun, 59045 Lille, France, 1Department of Chemistry, University of California, Irvine, CA 92697, USA and 2Department of Chemistry and Laboratory for Chemical and Biological Sciences, Georgia State University, Atlanta, GA 30303, USA
The antibiotic AT2433-B1 belongs to a therapeutically important class of antitumor agents. This natural product contains an indolocarbazole aglycone connected to a unique disaccharide consisting of a methoxyglucose and an amino sugar subunit, 2,4-dideoxy-4-methylamino-L-xylose. The configuration of the amino sugar distinguishes AT2433-B1 from its diastereoisomer iso-AT2433-B1. Here we have investigated the interaction of these two disaccharide indolocarbazole derivatives with different DNA sequences by means of DNase I footprinting and surface plasmon resonance (SPR). Accurate binding measurements performed at 4 and 25°C using the BIAcore SPR method revealed that AT2433-B1 binds considerably more tightly to a hairpin oligomer containing a [CG]4 block than to an oligomer with a central [AT]4 tract. The kinetic analysis shows that the antibiotic dissociates much more slowly from the GC sequence compared to the AT one. Preferential binding of AT2433-B1 to GC-rich sequences in DNA was independently confirmed by DNase I footprinting experiments performed with a 117 bp DNA restriction fragment. The specific binding sequence 5'-AACGCCAG identified from the footprints was then converted into a biotin-labeled DNA hairpin duplex and compound interactions with this specific sequence were characterized by high resolution BIAcore SPR experiments. Such a combined approach provided a detailed understanding of the molecular basis of DNA recognition. The discovery that the glycosyl antibiotic AT2433-B1 preferentially recognizes defined sequences offers novel opportunities for the future design of sequence-specific DNA-reading small molecules.
* To whom correspondence should be addressed. Tel: +33 320 16 92 18; Fax: +33 320 16 92 29; Email: bailly{at}lille.inserm.fr
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