Folding of phosphodiester-linked donor-acceptor oligomers into supramolecular nanotubes in water

Article abstract of DOI:10.1039/d1cc01064f

Inspired by the automated synthesis of DNA on a solid support, the electron-rich dialkoxynaphthalene (DAN) donor and the electron-deficient naphthalene-tetracarboxylic diimide (NDI) acceptor, amphiphilic foldamers have been synthesised from their respecti

Full text of DOI:10.1039/d1cc01064f

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Folding of phosphodiester-linked donor–acceptor
oligomers into supramolecular nanotubes in
water†
Cite this: Chem. Commun., 2021,
57, 4130
Received 25th February 2021,
Accepted 10th March 2021
Kevan Perez de Carvasal,a Nesrine Aissaoui,b Gerard Vergoten,c Gaetan Bellot,
b
´
´
´
¨
a
a
Jean-Jacques Vasseur,
Michael Smietana *a and François Morvan
*
DOI: 10.1039/d1cc01064f
rsc.li/chemcomm
Inspired by the automated synthesis of DNA on a solid support, the polymers or nanosheets. Along this line, foldamers built on electro-
electron-rich dialkoxynaphthalene (DAN) donor and the electron- static complementarity interactions (donor/acceptor) in water are
deficient naphthalene-tetracarboxylic diimide (NDI) acceptor, particularly important for the elaboration of novel materials, which
amphiphilic foldamers have been synthesised from their respec- are able to interfere with biological aqueous systems. Iverson and
tive phosphoramidite building blocks. The folding of the phos- co-workers have described the synthesis of self-complexing systems
phodiester-linked hexamer (DAN–NDI)3 revealed the formation of based on 1,5-dialkoxynaphthalene (DAN) and 1,8,4,5-naphthalen-
regular supramolecular nanotubes in water resulting from the self- etetracarboxylic diimide (NDI) motifs linked together with L-aspartic
assembly of multiple hexamers stabilized by donor/acceptor inter- acid17–19 or with mixed amino-acids to impart aqueous solubility.20
actions and the solvophobic effect.
Both aromatic units are stabilized in water by the solvophobic21
effect and donor–acceptor interactions known as charge-transfer
The tridimensional structure of supramolecular assemblies (CT).22–25 Alternatively, CT-based foldamers built from DAN and
plays an important role in their biophysical properties pyromellitic diimide units and tetra- or hexaethylene glycol linkers
and ultimately their functions regardless of whether they are complexed with alkali metals have also been reported.26
natural (DNA, proteins) or synthetic (molecular machines,
Herein, we present the synthesis and supramolecular assembly
polymers). In the quest for new folded structures and ultimately of new water-soluble DNA-inspired foldamers built from DAN and
in tuning their structural conformation to control their func- NDI moieties linked by negatively charged phosphodiesters leading
tion, foldamers have emerged as new tools for the development to the formation of nanotubes in water without the need of an
of biologically active synthetic molecules.1 Foldamers are linear organic solvent or any type of adjuvant (Fig. 1).
artificial oligomers that are able to fold into secondary struc-
The synthesis of these new structures started with the
tures stabilized by non-covalent forces including hydrogen preparation of NDI and DAN phosphoramidite building blocks
bonds, ionic interactions, p–p or s–p interactions, van der able to be directly assembled on an automated DNA synthesizer.
Waals forces or solvophobic driving force in water.2 While This chemical approach allows rapid access to various foldamer
DNA itself can be seen as a foldamer,3–5 several examples of sequences.27
folded DNA-inspired oligomers have been reported in the
The synthesis of NDI phosphoramidite 4 was proceeded by
literature. These structures are generally composed of repetitive reacting naphthalene dianhydride 1 with 3 equiv. of 3-amino-
aromatic6 polypyrene,7–13 polyanthracene14 or polyphenanthrene15 propanol to give the corresponding diimide 2 in 97% yield
units, except one built from perylenediimide–pyrene supramolecular (Scheme 1).28 Compound 2 was then treated with 1 equiv. of
interactions,16 which induce the formation of supratubular DMTrCl to give the monotritylated derivative 3 in a rather modest
yield (38%) due to the formation of a large amount of the ditritylated
a
´
´
Universite de Montpellier, CNRS, ENSCM, Institut des Biomolecules Max
Mousseron, Montpellier, France. E-mail: francois.morvan@umontpellier.fr,
michael.smietana@umontpellier.fr
b
c
´
Universite de Montpellier, INSERM, CNRS, Centre de Biochimie Structurale,
Montpellier, France
´
Universite de Lille, Inserm, INFINITE – U1286, Institut de Chimie Pharmaceutique
´
Albert Lespagnol (ICPAL), Faculte de Pharmacie, 3 rue du Professeur Laguesse,
Lille 59006, France
Fig. 1 Schematic representation of a nanotube built from the supra-
† Electronic supplementary information (ESI) available. See DOI: 10.1039/
molecular assembly of (DAN–NDI)3 foldamers.
d1cc01064f
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characterized by MALDI-TOF mass spectrometry (Table S1
and Fig. S1–S4, ESI†).
Compounds 9–11 were first analyzed using 1H-NMR spectro-
scopy (Fig. S5, ESI†). The DAN derivative 9 displays three types
of aromatic signals, 2 doublets and a triplet between 7.1 and
8.0 ppm, while the NDI derivative 10 displays only one single
aromatic signal at 8.75 ppm due to its symmetry. Compound 11
reveals multiple signals due to the asymmetry of the molecule
which leads to a polarization of the aromatics, thus confirming
their interaction. When the temperature was increased from
25 to 85 1C, a change of the folding state of 11 was observed.
At 25 1C, DAN and NDI aromatic protons exhibited a lower
chemical shift which indicates a strong shielding suggesting
that 11 is strongly folded on itself. When the temperature was
increased up to 85 1C, the strong deshielding of the aromatic
protons was progressively apparent. Indeed, the two doublets of
NDI aromatics moved closer together indicating a lower differentia-
tion of the hydrogen atoms as a result of fewer interactions between
DAN and NDI (Fig. S6, ESI†). In addition, the solution at high
temperature still exhibited pink color visible to the naked eye,
which is indicative of the presence of a CT-band. This study
demonstrates that increasing the temperature does not unfold
the structure completely.
Fig. 2 shows the UV/visible absorption spectra of the mathe-
matical sum of 9 and 10 (both at 99 mM) as well as 12 (33 mM) in
aqueous solution. The spectrum of 12 displayed an attenuation of
the absorption bands, a change of the two NDI intensity bands and
a 37% hypochromicity in comparison with the mathematical sum of
9 and 10. The charge transfer band is centered at 530 nm with a
molar extinction coefficient of 300 MÀ1 cmÀ1, but a higher concen-
tration is needed to clearly observe it (see Fig. 2, inset). The study
performed at 85 1C displayed a very similar spectrum with a slight
decrease of the CT band intensity demonstrating the strong folding
of 12 (Fig. S7, ESI†). All these changes are characteristic of a folded
structure and similar to those observed for previously reported DAN/
NDI foldamers.20,26,31 However, in contrast to DAN/NDI foldamers
built with amino acids,20 the current foldamer was very stable and
insensitive to temperature variations.
Scheme 1 Synthesis of NDI 4 and DAN 8 phosphoramidites.
side product. Finally, phosphitylation of
3 using N,
N-diisopropylcyanoethyl chlorophosphine gave NDI phos-
phoramidite 4 in 95% yield and an overall yield of 35%.
Concerning DNA phosphoramidite 8, any attempts to use
standard O-alkylation methods of 1,5-dihydroxynaphthalene 5
described in the literature were unsatisfactory and gave the
expected dialkyl derivative 6 in low yield (20 to 45%) along with
the formation of an oxidized side product.29,30 Hence the
alkylation was performed with 4 equiv. of 3-chloropropanol in
the presence of K2CO3 and KI in acetonitrile under reflux in an
atmosphere of argon for 18 h. Under these conditions, deriva-
tive 6 was isolated as a clean yellow-green powder in nearly
quantitative yield. Following the two-step protocol described
for the synthesis of 4, the DAN phosphoramidite 8 was obtained
in 46% overall yield.
With these phosphoramidites in hand, monomers of DAN 9
and NDI 10, as controls, as well as the alternated dimer 11 and
hexamer 12 were synthesized on a DNA synthesizer using
phosphoramidite chemistry and a custom-made propanediol
solid support (Scheme 2). To assure water solubility, an addi-
tional phosphodiester linkage was added at the end of each
sequence using a propanediol phosphoramidite unit 13.
After elongation, a methanolic ammonia treatment was
applied to deprotect and release compounds 9–12 from the
solid support. They were purified by C18 RP HPLC and
The fluorescence emission spectrum of compound 12 was
compared with the emission spectra of 9 and 10 (Fig. S8, ESI†).
Emissions of 9 and 10 concur with previous studies reporting
two high emissions of fluorescence at 330 and 345 nm18,32 for 9
(lex = 296 nm) and two high emissions of fluorescence at 390
Scheme 2 Synthesis of monomers 9 and 10, and foldamers 11 and 12.
SPOS: solid-phase oligonucleotide synthesis. SPOS conditions: (1) 3%
trichloroacetic acid (TCA) in CH2Cl2; (2) phosphoramidite derivative +
benzylthiotetrazole (BTT); (3) Ac2O, N-Me-imidazole, 2,6-lutidine; (4)
0.1 m I2 THF/H2O/pyridine.
Fig. 2 UV-vis spectra of oligomer 12 (33 mM, black curve) in water and the
mathematical sum of 9 and 10 (99 mM, red curve). Inset: Expanded region
of the charge-transfer band of 12 at 100 mM.
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Fig. 3 AFM images (height) at 45 mM in water after 1 d: (A) 9, (B) 10 and
(C) 9 + 10.
and 420 nm33 for 10 (lex = 363 nm). As expected, 12 exhibits a
very low emission of fluorescence at both excitation wave-
lengths due to the stable folding of the DAN and NDI units
with charge transfer that induces a very fast non-radiative
decay.32 The pink color which is characteristic of the CT
interaction observed for 12, associated with the important
hypochromicity observed in UV-vis spectra as well as the strong
decrease of fluorescence emission, confirmed the presence of a
folded structure.
The structures of the different compounds (9, 10, 9 + 10, 11
and 12) were imaged using atomic force microscopy (AFM) in
tapping-mode. Monomers alone dissolved in water (45 mM), for
1 day, form quite different structures. The DAN derivative 9
forms nanotubes (up to several hundreds of nm in length and
B2 nm in height) with some ramifications (Fig. 3A and Fig. S9,
ESI†), while the NDI derivative 10 forms vesicles (B30–200 nm
in diameter) (Fig. 3B and Fig. S10, ESI†). When both com-
pounds were mixed together in water, we observed strongly
ramified nanotubes with a length of several hundreds of nm
and a height of B1.5 nm (Fig. 3C and Fig. S11, ESI†).
Fig. 4 AFM images of 12 at 15 mM in water: (A) 1 d, height, (B) 1 d,
amplification, (C) 3 d, height, and (D) 7 d, height.
The formation of nanotubes for 12 was confirmed by TEM
imaging. A first study performed in water with the addition of
2% uranyl acetate solution for staining showed that the nano-
fibers folded on themselves to form a ball of twisted nanotubes
(Fig. S18, ESI†). Uranyl ions are known to bind with nucleic acid
phosphates and to induce aggregation.34 To reduce the for-
mation of related artifacts from uranyl acetate staining, we
adapted the buffer sample by adding 10 mM phosphate buffer
pH 7 and 10 mM NaCl. The TEM micrographs of the 15 mM
sample in this buffer showed micron scale one-dimensional
nanotubes similar to AFM images with fewer defects (Fig. 5 and
Fig. S19, ESI†).
According to the AFM study, foldamer 12 forms nanotubes
with an approximate thickness of 8 to 10 nm and lengths
of several micrometers. The molecular modelling study of
foldamer 12 shows a size of one unit of foldamer that is about
B2 nm long and B1.6 nm wide with a distance between two
aromatics of about 3.5 Å (Fig. 6A). The AFM images show
Dimer 11 was studied at a concentration of 45 mM after one
day in water. Only a very few nanotubes with a length up to
1 mm and heights between 0.5 and 1.2 nm were observed
(Fig. S12, ESI†).
collapsed tubes with a section around B16 nm and a height
11
¨
of B2 nm. In line with the work of Haner et al., the height,
Then, the folding of hexamer 12 was studied according to its
concentration and time of storage in solution. At 1 mM,
we observed only a few single nanotubes that are around
200–300 nm long and also ramified nanotubes that are up to
1 mm long with heights between 1 and 1.5 nm (Fig. S13, ESI†).
When the concentration was increased to 15 mM, more regular
and homogeneous nanotubes (100–400 nm long, 2 nm height)
were visualized (Fig. 4A and Fig. S14, ESI†) and at 30 mM even
longer nanotubes (200–500 nm long, 2 nm height) were
observed. Interestingly two nanotubes on top of each other
were also detected (Fig. S15, ESI†). The sample at a concen-
tration of 15 mM was then studied after 3 and 7 days to monitor
any changes over time (Fig. S16, ESI†). Nanotubes with a similar
width but a much higher length greater than 1 mm were
observed (Fig. 4C and D). Interestingly, when the 3 day old
solution was heated to 80 1C for 1 h, we observed the presence
of smaller structures, indicating a disassembly of the nano-
tubes upon heating (Fig. S17, ESI†). From the amplitude AFM
images, we could observe that this structure is a collapsed
nanotube with a thickness of 2 nm and a width of 16 nm
measured by AFM, corresponds to two layers of foldamers
indicating that the thickness of one layer is B1 nm corres-
ponding roughly to the length of aromatic motifs with a part of
the propyl arms. Hence, the tube has a perimeter of B32 nm
corresponding to 16 hexamers and a diameter of B10 nm was
also observed by TEM. The molecular modeling conducted with
these parameters with four turns is shown in Fig. 6B.
We calculated the longitudinal and lateral potential energies
of the interaction of two foldamers as À32.5 kcal molÀ1 and
À76.2 kcal molÀ1, respectively, corresponding to the energy
(Fig. 4B). These nanotubes have a similar structure to polyan-
Fig. 5 Negative-stained TEM images of 12 in 10 mM phosphate buffer
pH 7 and 10 mM NaCl, 15 mM 1 d.
11
¨
thracenes reported by Haner.
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Conflicts of interest
There are no conflicts to declare.
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