Conformational Diversity Versus Nucleic Acid Triplex
Stability, a Combinatorial Study*
Eloy
Bernal-Méndez and
Christian J.
Leumann
From the Departement für Chemie und Biochemie,
Universität Bern, Freiestrasse 3, CH-3012 Bern, Switzerland
The stability of a triple helix
formed between a DNA duplex and an incoming oligonucleotide strand
strongly depends on the solvent conditions and on intrinsic chemical
and conformational factors. Attempts to increase triple helix stability
in the past included chemical modification of the backbone, sugar ring,
and bases in the third strand. However, the predictive power of such modifications is still rather poor. We therefore developed a method that allows for rapid screening of conformationally diverse third strand oligonucleotides for triplex stability in the parallel pairing
motif to a given DNA double helix sequence. Combinatorial libraries of
oligonucleotides of the requisite (fixed) base composition and length
that vary in their sugar unit (ribose or deoxyribose) at each position
were generated. After affinity chromatography against their
corresponding immobilized DNA target duplex, utilizing a temperature
gradient as the selection criterion, the oligonucleotides forming the
most stable triple helices were selected and characterized by
physicochemical methods. Thus, a series of oligonucleotides were
identified that allowed us to define basic rules for triple helix
stability in this conformationally diverse system. It was found that
ribocytidines in the third strand increase triplex stability relative
to deoxyribocytidines independently of the neighboring bases and
position along the strand. However, remarkable sequence-dependent differences in stability were
found for (deoxy)thymidines and uridines.
*
This work was supported by the Swiss National Science
Foundation and Novartis AG, Basel.The costs of publication of this
article were defrayed in part by the
payment of page charges. The article must therefore be hereby marked
"advertisement" in
accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
To whom correspondence should be addressed. Tel.: 41-31-6314355;
Fax: 41-31-6313422; E-mail: leumann@ioc.unibe.ch.
Copyright © 2001 by The American Society for Biochemistry and Molecular Biology, Inc.
