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Nucleic Acids Research, 1994, Vol. 22, No. 3 479-484
© 1994

MOLECULAR BIOLOGY

Linkage structures strongly influence the binding cooperativity of DNA intercalators conjugated to triplex forming oligonucleotides

Frank M. Orson1,2,3,*, Berma M. Kinsey1,4 and W.Michael McShan1,2

1Veterans Affairs Medical Center Research Center on AIDS and HIV Infections Building 109, Room 226, VAMC, 2002 Holcombe, Houston, TX 77030, USA 2Department of Internal Medicine Building 109, Room 226, VAMC, 2002 Holcombe, Houston, TX 77030, USA 3Department of Microbiology and Immunology Building 109, Room 226, VAMC, 2002 Holcombe, Houston, TX 77030, USA 4Center for Biotechnology, Baylor College of Medicine Building 109, Room 226, VAMC, 2002 Holcombe, Houston, TX 77030, USA

*To whom correspondence should be addressed at: Building 109, Room 226, Veterans Affairs Medical Center, 2000 Holcombe Boulevard, Houston, TX 77030, USA

Received August 23, 1993. Accepted November 29, 1993.

Conjugation of DNA intercalators to triple heiix forming oiigodeoxynucieotides (ODN's) can enhance ODN binding properties and consequentiy their potentiai abiilty to moduiate gene expression. To test the hypothesis that iinkage structure couid strongiy Infiuence the binding enhancement of intercaiator conjugation with triplex forming ODN's, we have used a modei system to investigate binding avidity of short oiigomers conjugated to DNA intercaiators through various linkages. Using a dA10·T10 target sequence imbedded in a 20 bp duplex, binding avidities of a T10 ODN joined to the DNA intercalator 6,9-diamino, 3-methoxy acridine (DAMA) by 8 different 5' linkages were measured using an electrophoretic mobiiity shift assay. Afthough unmodified T10 has a very limited capacity for stabie binding under these conditions (apparent Kd >250 µM at 4°C), conjugation to DAMA using tiexible linkers of certain lengths and chemicai compositions greatly enhanced binding (Kd of 1 µM at 4°C). Other linkers, however, modestiy enhanced binding or had no effect on binding at au. Thus, the length, flexibility, and chemicai composition of iinker structures au substantialiy influence intercalator conjugated oilgodeoxynucleotide binding avidity.


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