Hairpin mimics with phenanthroline- and bipyridine-derived linkers

Article abstract of DOI:10.1080/15257770701508026

The synthesis and structural stabilities of modified oligonucleotide hairpins containing phenanthroline- and bipyridine-modified loops is reported. Phenanthroline (phen) and bipyridine (bipy) building blocks were synthesized, incorporated into DNA-oligonucleotides, and analyzed by thermal denaturation experiments. The so modified oligomers were found to form stable hairpin structures. Tm values were not affected by divalent transition metals. Copyright Taylor & Francis Group, LLC.

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Hairpin Mimics with Phenanthroline- and
Bipyridine-Derived Linkers
Gapian Bianké a & Robert Häner a
a Department of Chemistry , Biochemistry, University of Bern , Bern,
Switzerland
Published online: 10 Dec 2007.
To cite this article: Gapian Bianké & Robert Häner (2007) Hairpin Mimics with Phenanthroline-
and Bipyridine-Derived Linkers, Nucleosides, Nucleotides and Nucleic Acids, 26:8-9, 949-952, DOI:
10.1080/15257770701508026
To link to this article: http://dx.doi.org/10.1080/15257770701508026
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Nucleosides, Nucleotides, and Nucleic Acids, 26:949–952, 2007
Copyright ꢀ Taylor & Francis Group, LLC
C
ISSN: 1525-7770 print / 1532-2335 online
DOI: 10.1080/15257770701508026
HAIRPIN MIMICS WITH PHENANTHROLINE- AND
BIPYRIDINE-DERIVED LINKERS
Gapian Bianke´ and Robert Ha¨ner
Department of Chemistry and Biochemistry,
2
University of Bern, Bern, Switzerland
The synthesis and structural stabilities of modified oligonucleotide hairpins containing
2
phenanthroline- and bipyridine-modified loops is reported. Phenanthroline (phen) and bipyridine
(bipy) building blocks were synthesized, incorporated into DNA-oligonucleotides, and analyzed by
thermal denaturation experiments. The so modified oligomers were found to form stable hairpin
structures. Tm values were not affected by divalent transition metals.
Keywords oligonucleotide
hairpins;
phenanthroline;
bipyridine;
thermal
denaturation.
Metal-coordinating oligonucleotide conjugates combine the ability of se-
quence specific DNA or RNA recognition with the function of metal coor-
dination through appropriate ligands. This class of bioconjugates is actively
investigated due to their importance in the field of artificial nucleases,[1−3]
the self assembly of metals arrays,[4,5] electron transfer,[6,7] and as sequence
specific probes and sensors.[8−10] We have endeavored in the design, synthe-
sis, and structural investigation of metal-coordinating DNA hairpin mimics.
The hairpin is a secondary structural motif of nucleic acid which is most
commonly found in all biological systems.[11] Due to its important role in
the structural and functional behavior of nucleic acid, it has been the sub-
ject of numerous studies. Thus, several nonnucleosidic scaffolds such as
oligo ethylene glycol linkers,[12] phenanthrene,[13] stilbene, naphthalene,
and diphenylacetylene[14] have been reported as loop replacements in DNA
hairpins, as well as metal-bridged15 hairpins and metallo-salen[16] derived
DNA hairpins. Metal coordinating ligands as loop replacements offer the
possibility of metal-coordination in addition to serving as a structural scaf-
fold. Previously, we have reported that the replacement of terpyridine in
DNA loop shows a strong improvement of hairpin stability in the absence of
metal in comparison to the dT4- and dA4-loop.[17] We extended this work
Address correspondence to Robert Ha¨ner, Department of Chemistry and Biochemistry, University
of Bern, Freiestrasse 3, CH-3012 Bern, Switzerland. E-mail: robert.haener@ioc.unibe.ch
949

950
G. Bianke´ and R. Ha¨ner
FIGURE 1 Illustration of phenanthroline- and bipyridine-modified hairpin structures.
to different oligopyridine ligands that are known to bind to transition metal
ions. Here, we report the synthesis and properties of a bipyridine and a
phenanthroline based metal-coordinating hairpin mimic (Figure 1).
Following the known synthesis of bipyridine derivatives,[18,19] 6,6ꢁ-
bis(hydroxymethyl)-[2,2ꢁ]-bipyridine (1) was prepared. Mono-protected
compound 2 was obtained after reaction of 1 with 4,4ꢁ-dimethoxytrityl chlo-
ride (DMT-Cl). Compound 2 was further converted into the correspond-
ing phosphoramidite 3 by conventional phosphitylation (Scheme 1). The
dimethyl ester of [1,10]phenanthroline derivative 4 was synthesized accord-
ing to the described procedures.[20,21] Amides 5a and 5b, containing a 2-
and 3-carbon linker were obtained by reaction of 4 with 2-aminoethanol
SCHEME 1 (a) 4,4ꢁ-Dimethoxytrityl chloride, pyridine, r.t. 24 hours; (b) (iPr2N)2P(OCH2CH2CN),
N,N-diisopropylammonium 1H-tetrazolide, CH2Cl2, r.t, 1 h. DMT: 4,4ꢁ-dimethoxytrityl; PAM: N ,N ꢁ-
diisopropyl-2-cyanoethyl phosphoramidite.
SCHEME 2 (a) 2-Aminoethanol (or 3-aminopropanol), reflux, 1 hour; (b) ) 4,4ꢁ-dimethoxytrityl chlo-
ride, pyridine, r.t. overnight; (c) (iPr2N)2P(OCH2CH2CN), N,N-diisopropylammonium 1H-tetrazolide,
CH2Cl2, r.t. 90 minutes.

Hairpin Mimics
951
TABLE 1 Phenanthroline- and bipyridine-containing hairpins 8–10 and control oligonucleotides
11-13 used in this study and Tm values obtained by thermal denaturation experiments. Conditions:
1.5 µM oligomer concentration, 10 mM phosphate buffer, 100 mM NaCl, pH = 7.5; temperature
gradient: 0.5◦C/minute. Melting temperatures were determined from the maximum of the first
derivative of the melting curve (A260 against temperature); exptl. error 0.5◦C. Abbreviations
(C2) and (C3) refer to (CH2)2- and (CH2)3-linkers, respectively.
Tm(◦C)
Oligomers
Sequences
(−)
Co(II)
Zn(II)
Cu(II)
a
8
9
10
11
12
13
5ꢁ-GCAATTGC-phen(C2)-GCAATTGC-3ꢁ
5ꢁ- GCAATTGC-phen(C3)-GCAATTGC-3ꢁ
5ꢁ- GCAATTGC-bipy-GCAATTGC-3ꢁ
5ꢁ- GCAATTGC- 3ꢁ
64.0
63.0
62.1
29.0
73.6
74.6
65.4
62.7
64.5
63.5
—
—
a
a
a
—
—
63.0
a
a
a
—
—
—
a
a
a
5ꢁ- GCAATTGC-AAAA-GCAATTGC-3ꢁ
5ꢁ- GCAATTGC-TTTT-GCAATTGC-3ꢁ
—
—
—
74.4
74.6
74.8
a
Values not determined.
and 3-aminopropanol, respectively. Mono-protection of 5a and 5b with
DMT-Cl afforded the corresponding compounds 6a and 6b which were fur-
ther transformed into desired phenanthroline phosphoramidites 7a and 7b
(Scheme 2).
The syntheses of the phenanthroline- and bipyridine-modified
oligomers 8, 9, and 10 were performed by automated oligonucleotide syn-
thesis (Table 1). Oligonucleotides 11, 12, and 13 were prepared for control
purposes.
The stability of the oligonucleotides in the presence and absence of
different metals was analysed by thermal denaturation experiments. The
Tm (melting temperature) values obtained are shown in Table 1. Tm val-
ues were independent of the oligomer concentration (0.5–5 µM) demon-
strating that the observed process is monomolecular (data not shown). The
phenanthroline-modified hairpins 8 and 9 as well as the bipyridine-modified
FIGURE 2 Influence of different metals on the thermal denaturation curves of the phenanthroline-
modified hairpins 8 (a) and 9 (b) as well as the bipyridine-modified hairpin 10 (c). Conditions: 1.5 µM
oligomer concentration, 10 mM NaH2PO4, 100 mM NaCl, pH = 7.5.

952
G. Bianke´ and R. Ha¨ner
hairpin 10 had lower Tm’s (64.0, 63.0, and 62.1◦C, respectively) than the
corresponding dA4- or dT4-hairpins in the absence of polyvalent cations.
This is in contrast to previous reports,[12−14] for which generally a substan-
tial increase in the Tm was observed. Most importantly, no change in the
Tm-values was observed with phenanthroline- and bipyridine-modified hair-
pins upon addition of metals, such as Co(II), Zn(II) or Cu(II) as shown in
Figure 2.[22]
CONCLUSIONS
Phenanthroline and bipyridine phosphoramidite building blocks were
synthesized and incorporated into self complementary oligonucleotides.
The so modified oligomers were found to form stable hairpin structures.
In comparison to the analogous control hairpins containing either a dA4-
or dT4-hairpin loop, all modified structures were found to be less stable.
Moreover, the hairpin mimics were not sensitive to the presence of divalent
metals, i.e., Co(II) or Zn(II) in the case of the phenanthroline- and Cu(II)
in the case of the bipyridine-modified hairpin.
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