Design of sequence specific DNA binding molecules: bis(distamycin)phenoxazone

Abstract

One approach to the design of sequence specific DNA binding molecules that read large sequences of right-handed double helical DNA is to couple DNA binding units of similar or diverse base pair specificities. The natural product distamycin is a tris-N-methylpyrrolecaboxamide tripetide that binds in the minor groove of DNA at A•T rich sequences that are five base pairs in size. The natural product actinomycin D which consists of an aromatic phenoxazone coupled to two identical pentapeptide lactones is an intercalator which binds four base pairs with a preference for 5'-NGCN-3' sequences. The synthesis of a hybrid distamycin and actinomycin, bis(distamycin)phenoxazone is described where the cyclic pentapeptides of actinomycin are replaced by the tripeptide of distamycin. Using the affinity cleaving method we find that bis(EDTA-distamycin)phenoxazone (BEDP) binds the sequence 5'-TATAGGTTAA-3' consistent with simultaneous binding of the tripeptides flanking the intercalator. At other sites the results indicate that the tripeptides do not bind simultaneously. One interpretation of the data suggests that the distortion of the DNA sequences flanking the phenoxazone intercalation site may be sequence specific and thus prevent binding of both tripeptide groover binders.

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