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Nucleic Acids Research, 1994, Vol. 22, No. 19 3977-3982
© 1994

RNA

Comparison of antiparallel A-AT and T-AT triplets within an alternate strand DNA triple helix

Elinor Washbrook and Keith R. Fox*

Department of Physiology and Pharmacology, University of Southampton Bassett Crescent East, Southampton SO9 3TU, UK

*To whom correspondence should be addressed

Received May 11, 1994. Revised August 15, 1994. Accepted August 15, 1994.

We have examined the formation of alternate strand triple-helices at the target sequence A11(TC)6-(GA)6T11 using the oligonucleotides T11(AG)6 and T11(TG)6, by DNase I footprinting. These third strands were designed so as to form parallel T-AT triplets together with antiparallel G-GC and A-AT or T-AT triplets. We find that, although both oligonucleotides yield clear footprints at similar concentrations (0.3 µM) in the presence of manganese, only T11(TG)6 forms a stable complex in magnesium-containing buffers, albeit at a higher concentration (10–30 µM). Examination of the interaction of (AG)6 and (TG)6 with half the target site confirmed that the complex containing A-AT triplets was only stable in the presence of manganese. In contrast no binding of (TG)6 was detected in the presence of either metal ion, suggesting that the reverse-Hoogsteen T-AT triplet is less stable that GGC. We suggest that, within the context of GGC triplets, the rank order of antiparallel triplet stability is A·AT(Mn2+) > T.AT(Mn2+) > T.AT (Mg2+) > A.AT (Mg2+). Third strands containing a single base substitution in the centre of either the parallel or antiparallel portion showed a (10-fold) weaker interaction in manganese-containing buffers, and no interaction in the presence of magnesium.


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