A conjoined thienopyrrole oligomer formed by using DNA as a molecular guide

Article abstract of DOI:10.1021/ol801137t

(Figure Presented) A thienopyrrole oligomer conjoined to DNA was prepared by means of a templated synthesis protocol. The oligomer was formed by reaction, initiated with HRP/H₂O₂, of thieno[3,2-b]pyrrole monomers attached to cytosine bases. The thienopyrrole oligomer was characterized spectroscopically.

Full text of DOI:10.1021/ol801137t

ORGANIC
LETTERS
2008
Vol. 10, No. 17
3657-3660
A Conjoined Thienopyrrole Oligomer
Formed by Using DNA as a Molecular
Guide
Selvi Srinivasan and Gary B. Schuster*
School of Chemistry and Biochemistry, Georgia Institute of Technology, Atlanta,
Georgia, 30332
schuster@gatech.edu
Received May 28, 2008
ABSTRACT
A thienopyrrole oligomer conjoined to DNA was prepared by means of a templated synthesis protocol. The oligomer was formed by reaction,
initiated with HRP/H2O2, of thieno[3,2-b]pyrrole monomers attached to cytosine bases. The thienopyrrole oligomer was characterized
spectroscopically.
The creation and manipulation of well-defined nanoscale
structures with high precision depends ultimately on the
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10.1021/ol801137t CCC: $40.75
Published on Web 08/08/2008
2008 American Chemical Society

Extensive investigation of the properties of DNA has
revealed that its electrical characteristics are not well-suited
for functions that require the rapid and efficient transport of
charge over long distances.15-18 The application of DNA to
molecular electronics as a “wire” therefore requires its major
modification9,19-22 or an appropriate restructuring of the
DNA itself23,24 while continuing to take advantage of its self-
organizing and self-recognizing properties. An example of
this approach is metallization procedures9-11 that form
conductive structures along a path originally defined by the
DNA. Similarly, the fabrication of DNA attached to semi-
conductor nanoparticles19,25 or to conducting polymers26-28
offers considerable promise.
Conducting polymers such as polypyrrole and polya-
niline26-29 have been synthesized by taking advantage of
electrostatic interactions to organize their positively charged
monomers along the negatively charged phosphate backbone
of DNA. This strategy is easy to execute, but it does not
allow for specific attachment of the conducting polymer and
does not take advantage of the sequence information inherent
in DNA. We have been examining the possibility of forming
DNA-linked conducting polymers that are attached co-
valently at specified bases, thus combining the templating
and scaffolding roles of DNA. As an initial step toward this
goal, we recently showed that either horseradish peroxidase
(HRP) under mild oxidizing conditions or electrochemical
methods can be employed to form polyaniline (PANI) from
aniline derivatives linked to the bases of an oligonucleotide
without destroying the duplex.30,31 In this communication,
we report the extension of this strategy to a heterocyclic
monomer. Thieno[3,2-b]pyrrole monomers attached to cy-
tosine bases through a flexible three-carbon chain were
Figure 1. Schematic representation of the DNA oligomers used in
this work. The symbol X stands for a cytosine nucleobase having
a covalently attached thienopyrrole monomer prepared by postsyn-
thetic modification of the oligonucleotide, as is shown.
oxidized, and optical absorption experiments indicated that
this reaction results in the formation of a conducting polymer.
Thienopyrrole is an electron-rich heteroaromatic bicyclic
compound containing pyrrole and thiophene units fused
together. This compound is more easily oxidized (Eox ) 0.6
V vs Ag/Ag+) than thiophene as a consequence of its greater
π-electron delocalization. It has been predicted from ab initio
theoretical calculations that poly(thienopyrrole) has a pre-
dominant R-R′ linkage between the monomer units. Simi-
larly, poly(thienopyrrole) is expected to have good conduc-
tivity and be easily transformed between its oxidized and
reduced states,32 making it an ideal candidate for use as a
nanowire conjoined to DNA. Moreover, preparation of
oligonucleotides having thienopyrole groups on cytosines
seemed straightforward and in fact was readily accomplished.
The DNA oligomers used in the current study are shown
in Figure 1 where X represents the modified cytosine bearing
the thienopyrrole moiety. They were prepared by postsyn-
thetic modification of the appropriate oligonucleotides.
1-(3-Aminopropyl)thieno[3,2-b]pyrrole (TP) was prepared
from thieno[3,2-b]pyrrole (5), which in turn was effi-
ciently obtained from thiophene-2-carboxaldehyde (1) by
Hemetsberger-Knittel reaction methodology (see Scheme
1).33 DNA(1) and DNA(2) are sequences with six and one
TP units, respectively. The arrangement of alternating X and
T nucleotides of DNA(1) was selected on the basis of
molecular mechanics calculations so that the monomers were
properly spaced for reaction. DNA(3) is a normal, unmodified
sequence used in control experiments. DNA(4) and DNA(5)
are the complementary sequences to the above oligonucle-
otides. All DNA oligomers were purified by reverse-phase
HPLC, and the structures were confirmed by optical and ESI
mass spectrometry.
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Duplex DNA oligomers were prepared by hybridization
of the appropriate single strands and probed by examination
of their thermal melting behavior, first to confirm that
duplexes are formed, but primarily to ensure that they are
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3658
Org. Lett., Vol. 10, No. 17, 2008

Scheme 1. Synthetic Route Used for the Preparation of
1-(3-Aminopropyl)thieno[3,2-b]pyrrole
Figure 2
. Absorption spectra of DNA(1/4), DNA(2/5), and DNA(1)
(5 µM) before and after treatment with HRP/H2O2.
stable at the temperatures where the oligomerization reactions
are carried out. The duplex formed from DNA(3/4), which
contains no modified cytosines, was observed to melt (Tm)
at 72 °C, whereas DNA(1/4), which contains six modified
cytosines, has Tm ) 49 °C. Thus, modification of the DNA
by incorporation of TP monomers results in a considerable
stability decrease, but DNA(1/4) clearly exhibits the neces-
sary stability for reaction with HRP at room temperature.
We also determined the circular dichroism (CD) spectrum
of DNA(1/4), which exhibits bands between 200 and 300
nm that resemble that of normal B-form DNA.34 These
experiments indicate that incorporation of the six TP units
in DNA(1/4) does not destroy or overly distort the duplex
structure.
It is well established that aniline-like monomers covalently
attached to DNA can be oligomerized under oxidizing
conditions.30,31 The reaction of DNA(1/4) with H2O2 and
HRP in citrate buffer at pH 4.5 was monitored by UV-vis
spectroscopic measurements taken at 15-min intervals. As
is shown in Figure 2, the reaction of DNA(1/4) under these
conditions results in the growth of two new absorption bands,
one at 547 nm and the other at 806 nm. These absorption
bands are readily apparent within 15 min of the start of the
reaction, reach a maximum intensity within 1 h, and then
remain unaltered for several hours. No such absorption
changes are observed when DNA(3/4), which does not
contain a thieno[3,2-b]pyrrole unit, is subjected to these
reaction conditions.
DNA(2/5) contains only one TP-containing modified
nucleobase. It was examined in a control experiment intended
to assess whether the spectral changes observed in the
reaction of DNA(1/4) are the result of reactions between
several monomer units or an oxidative process involving a
single thieno[3,2-b]pyrrole group. Treatment of DNA(2/5)
with H2O2 and HRP does not result in the appearance of
significant absorption bands at 547 or 806 nm. Also, the
reaction of the single strand DNA(1) with HRP/H2O2 results
Figure 3. Agarose gel of duplex DNA(1/4) before (A) and after
(B) treatment with HRP/H2O2.
in a product having an absorption band only at ca. 520 nm
(see Figure 2), which is distinctly different from that formed
in the reaction of DNA(1/4) duplex.
Melting temperature experiments show that after reaction
with HRP/H2O2 the Tm of DNA(1/4) is broad with a
maximum at 46 °C, which indicates that formation of the
conjoined oligomer maintains a duplex structure but that it
is distorted. However, attempted hybridization of the product
formed from oxidation of single strand DNA(1) with its
complement (DNA(4)) did not give a duplex. This finding
supports the conclusion that the treatment of the single strand
and the duplex with HRP and H2O2 give different products.
Finally, agarose gel electrophoresis analysis (see Figure
3 and Supporting Information) of DNA(1/4) after its reaction
with HRP/H2O2 shows that there has been no cross-linking
of the DNA strands or branching of TP monomers in the
formation of oligomer. These results indicate that oxidation
of duplex DNA(1/4) with HRP/H2O2 results in the formation
(34) Gray, D. M.; Hung, S.-H.; Johnson, K. H. Methods Enzymol. 1995,
246, 19–36.
Org. Lett., Vol. 10, No. 17, 2008
3659

of a short-chain, linear thienopyrrole oligomer that has the
absorption spectrum expected for the conducting polymer.
The UV-vis absorption spectrum of poly(thienopyrrole)
has not been previously reported. We prepared this com-
pound by standard procedures35,36 in order to confirm that
the absorption changes observed upon oxidation of DNA(1/
4) are indeed characteristic of formation of the expected
conjoined oligomer. The oxidation of thieno[3,2-b]pyrrole
(5) in acetonitrile with FeCl3 results in the formation of a
polymer having optical absorption bands at 546 and 797 nm
(see Figure 4). These bands are similar to those that result
from the reaction of DNA(1/4) with HRP/H2O2, which
indicates that the latter process gives a poly(thienopyrrole).37
In this work we have successfully demonstrated the DNA-
directed synthesis of thienopyrrole oligomers covalently
tethered to DNA. This DNA conjoined oligomer system was
formed by taking advantage of the highly specific self-
assembly properties of DNA. It shows that the formation of
DNA-conjoined polymers is a general phenomenon not
limited to simple derivatives of aniline. This finding signifi-
cantly expands the number of possible monomers that may
be used to create conjoined oligomers along a pathway
defined by DNA. Further studies are underway that include
extending the chain length of the oligomer and evaluating
the ability of a single polymer molecule to function as an
electical conductor.
Figure 4. Absorption spectra representing polythienopyrrole forma-
tion in acetonitrile with FeCl3 oxidant. The UV region is not shown
because the FeCl3 absorption obscures the region.
Acknowledgment. We thank the Vassar Woolley Founda-
tion for financial support, and Dr. Bhaskar Datta and Dr.
Joshy Joseph (School of Chemistry and Biochemistry,
Georgia Institute of Technology) for assisting with DNA
syntheses and for helpful discussions.
(35) Li, H.; Lambert, C.; Stahl, R. Macromolecules 2006, 39, 2049–
Supporting Information Available: Procedures for the
synthesis of TP monomer, TP modified DNA oligomers and
agarose gel electrophoresis, ESI mass spectrometric char-
acterization of modified DNA sequences, and thermal melting
curves and circular dichroic spectra of DNA(1/4). This
material is available free of charge via the Internet at
http://pubs.acs.org.
2055
.
(36) Machida, S.; Miyata, S.; Techagumpuch, A. Synth. Met. 1989, 31,
311–318
.
(37) Differences in the absorption maxima of the oligomer bands shown
in Figures 2 and 4 likely result because the former is recorded in citrate
buffer solution, the oligomer is constrained by its covalent attatchment to
DNA, and its chain length is a maximum of 6 monomer units. Whereas the
polymer whose spectrum is shown in Figure 4 is dissolved in acetonitrile
solution, its structure is unconstrained and it is composed of a variable
number of monomer units. The changes in solvent, structure, and length
may be responsible for the 10 nm shift in the observed absorption maxima.
OL801137T
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Org. Lett., Vol. 10, No. 17, 2008