Self-Assembly of a Guanosine Derivative To Form Nanostructures and Transmembrane Channels

Article abstract of DOI:10.1002/chem.201800205

We herein report the self-assembly of a lipophilic bromoguanosine derivative (G1) in homogeneous solution, in the solid state and in planar bilayer membranes. The self-assembly of G1, driven by H-bonding and π–π stacking interactions can form different nano-structures depending on incubation time. The G1 nanostructure is able to bind a bioactive dye like Rose Bengal. In crystal state, it shows ribbon type H-bonding pattern and exhibits birefringence in polarized light. And further, the self-assembled nanostructure of G1 can form discrete transmembrane ion channels in lipid bilayer membranes, enabling passage of potassium ions.

Full text of DOI:10.1002/chem.201800205

A Journal of
Accepted Article
Title: Self-assembly of a Guanosine Derivative to Form Nanostructures
and Transmembrane Channels
Authors: Jyotirmayee Dash
This manuscript has been accepted after peer review and appears as an
Accepted Article online prior to editing, proofing, and formal publication
of the final Version of Record (VoR). This work is currently citable by
using the Digital Object Identifier (DOI) given below. The VoR will be
published online in Early View as soon as possible and may be different
to this Accepted Article as a result of editing. Readers should obtain
the VoR from the journal website shown below when it is published
to ensure accuracy of information. The authors are responsible for the
content of this Accepted Article.
To be cited as: Chem. Eur. J. 10.1002/chem.201800205
Link to VoR: http://dx.doi.org/10.1002/chem.201800205
Supported by

Chemistry - A European Journal
10.1002/chem.201800205
COMMUNICATION
Self-assembly of a Guanosine Derivative to Form Nanostructures
and Transmembrane Channels
[
a]
[a]
[a]
[b]
[b]
Rabindra Nath Das, Y. Pavan Kumar, S. Arun Kumar, Ole Mathis Schütte, Claudia Steinem,
[
a]
Jyotirmayee Dash *
Abstract: We herein report the self-assembly of
a lipophilic
bromoguanosine derivative (G1) in homogeneous solution, in the
solid state, and in planar bilayer membrane. The self-assembly of
G1, driven by H-bonding and - stacking interactions can form
different nano-structures depending on incubation time. The G1
nanostructure is able to bind a bioactive dye like Rose Bengal. In
crystal state, it shows ribbon type H-bonding pattern and exhibits
birefringence in polarized light. And further, the self-assembled
nanostructure of G1 can form discrete transmembrane ion channels
in the biological membrane, enabling transportation of potassium
ions.
Figure 1. Guanosine and its derivatives.
The images showed that the TBDMS protected bromoguanosine,
G1 formed circular honeycomb type supramolecular structures
after 1 h incubation (Figure 2 a, b).[6] The average height of the
ring was determined to be 30 nm and the average width was
Guanosine derivatives, owing to their self-assembling properties,
have been used as building blocks for fabricating well-defined
nanostructures, hydrogels
and synthetic ion channels.[1]
125 nm. The images also showed that some of these rings were
Guanosine derivatives can self-assemble using Hoogsteen type
hydrogen bonding and π-π stacking interactions to generate
tunable three-dimensional highly ordered structures.[1] Lipophilic
guanosine derivatives, depending on the substituents and the
experimental conditions, are able to form supramolecular
fused together forming dumbbell shaped loops. Interestingly,
incubating a solution of G1 for 4 days, a network of G-wires
interconnected at few nodes was observed (Figure 2 c, d). AFM
and TEM images highlight the dependence of self-assembly of
G1 on incubation time. These results indicate that the ring or
dumbbell shaped self-assembled structures of G1 open up and
join end-to-end to form longer and thinner G-wire architectures
with time.
[
2]
architectures like ribbons, sheets and helices. In the
presence of metal ions, guanosine derivatives self-assemble
to form macrocyclic G-quartet,[1a] which is a prime building
block of biologically relevant four stranded DNA and RNA
secondary structures.[3] The G-quartet like assembly has been
demonstrated to incorporate aromatic dyes and drug molecules;
useful for industrial and biomedicinal applications.[1] It has been
reported that 8-substituted guanosine derivatives form stable
self-assembled structures[4,5] as C-8 modification causes the
nucleobase to preferentially adopt the syn-conformation and
enables Hoogsteen type H-bonding. In our previous report, we
have used
a combination of guanosine 1 and 8-bromo
[
5]
guanosine 2 to prepare stable hydrogels. We herein report the
self-assembling and ion transportation properties of
a 8-
bromosubstituted lipophilic guanosine derivative G1.
The lipophilic tert-butyldimethylsilyl (TBDMS) protected
bromo guanosine derivative G1 was prepared from guanosine
nucleoside 1 (Figure 1, Scheme S1, see Supporting Information,
S.I.). The self-assembling property of G1 on solid surfaces was
studied using AFM and TEM imaging.
[
[
a]
b]
R. N. Das, Y. P. Kumar, S. A. Kumar, Prof. Dr. J. Dash
Department of Organic Chemistry
Indian Association for the Cultivation of Science
Jadavpur Kolkata-700032, India
Figure 2. AFM and TEM images of G1 after 1 h (a, b); 4 days (c, d) incubation
E-mail: ocjd@iacs.res.in
(scale bar 400 nm).
O. M. Schütte, Prof. Dr. C. Steinem
Institute for Organic and Biomolecular Chemistry
Georg August University Göttingen
Tammannstr. 2, 37077 Göttingen, Germany
email: csteine@gwdg.de
The solid-state conformation of G1 was studied using single
crystal X-ray analysis.[7] The structure analysis reveals that it
forms H-bonded 1D chain like structures in which each
guanosine is connected through four Hoogsteen type H-bonding
Supporting information for this article is given via a link at the end of
the document.
This article is protected by copyright. All rights reserved.

Chemistry - A European Journal
10.1002/chem.201800205
COMMUNICATION
with four-neighbouring guanosine motifs (Figure 3). In these
extended H-bonded layers, the amino (N2) hydrogen of a
guanosine motif (A) H-bonds with N7 of another guanosine motif
(
B); the imino hydrogen of A is further H-bonded with the O6 of
guanosine motif (C). In a similar fashion, the O6 and N7 of A are
H-bonded with imino and amino hydrogens of guanosine motifs
D and E, respectively leading to the formation of a ribbon type
nanowire.[2a,e] The extended H-bonded networks of G1 further
held together through stacking interactions of the aromatic rings.
It has been reported that guanine crystals are responsible for
amazing color in animals e.g. fish skin, widow spider etc. due to
their extremely high refractive index.[8] We observed that G1
crystals exhibited strong birefringence upon exposure to
[5]
polarized light (Figure 3c). This result indicates that G1 crystal
is anisotropic in nature due to the difference in the refractive
indices (n) along two different parallel H-bonded crystallographic
directions. The densely stacked H-bonded layers present in the
crystal structure exhibiting birefringence would be of great
Figure 4. 1H- NMR spectra of G1 in CDCl
5 °C).
3
at variable temperatures (-40 °C-
2
It has been demonstrated that guanosine derived hydrogels
can find useful applications in drug-delivery as well as optical
and organic electronic systems as the supramolecular assembly
interest in different applications like data storage, development
of optical devices and bio-imaging.[9]
can bind bioactive aromatic dyes using - stacking and
electrostatic interactions.[
5,11]
To gain insights into the binding
property of the G1 assembled nano-construct, we studied the
emission and steady state anisotropy of a bioactive dye rose
bengal in the presence of G1. The fluorescence intensity of rose
bengal was significantly decreased suggesting its incorporation
into the G1 nano-construct (Figure 5a). We also observed a
remarkable enhancement in the fluorescence anisotropy of rose
bengal in the presence of G1 and that further confirms the
binding of the dye molecules into G1 network (Figure S2). AFM
imaging was performed to investigate the morphological
changes of G1 assembled nanostructures after the
encapsulation of rose bengal. The AFM images showed the
accumulation of dyes in G1 network (Figure 5b).
Figure 3. (a) Crystal structure of G1, (b) H-bonding pattern in G1, (c) G1
crystals exhibiting birefringence.
The self-assembly of G1 was then studied in solution state
1
(
CDCl
3
) by using variable temperature (VT) NMR (Figure 4). H
NMR spectra of G1 were recorded at various temperatures (-40
to 25 °C) to probe the intermolecular H-bonding (N1-O6 and N2-
N7). At room temperature (25 °C), G1 showed a strong broad
peak at 6.52 ppm for the –N(2)H
of protons. By decreasing the temperature (-40 °C), two broad
peaks at 8.82 and 4.95 ppm were observed for –N(2)H protons.
At low temperature, the exchange rate of the H-bonded protons
became slow enabling discrimination of –N(2)H protons. In
2
protons, due to fast exchange
2
2
addition, a relatively sharp peak at 12.12 ppm for –N(1)H was
also observed by lowering the temperature. These results
indicate the formation of a H-bonded network of G1 in solution
Figure 5. (a) Fluorescence spectra of rose bengal (10 M) with addition of G1,
(b) AFM image of G1 after the encapsulation of rose bengal (scale bar 400
nm)
+
state. The self-assembly of G1 in the presence of K was also
studied by using NMR spectroscopy.[10] 1H NMR spectra of G1 in
the presence of K-picrate showed a peak at 12.0 ppm indicating
the involvement of imino–N(1)H proton in the H-bonding to form
G-quartet like arrangement (Figure S1).
Guanosine derivatives have also been used as building
blocks for synthetic ion channels.[1f, 12-15] The Davis group
reported that a lipophilic guanosine derivatives can assemble
into a unimolecular G-quadruplex structure that can transport
Na+ ions across a phospholipid bilayer.[12] Matile and Kato
groups reported that a folic acid derivative containing similar H-
bonding motif as in guanine can self-assemble to form ion
channels.[13] Subsequently, diguanosine derivatives containing
different covalent linkers have been demonstrated to form
membrane spanning channels by self-assembly.[
14,15]
In recent
years, there is an emerging interest in the development of
This article is protected by copyright. All rights reserved.

Chemistry - A European Journal
10.1002/chem.201800205
COMMUNICATION
synthetic ion channels that can mimic the structural and
functional aspects of natural ion channels.[16]
Synthetic
channels can be useful for a diversity of applications like
therapeutics[17] and sensors.[18]
Small molecules, peptides,
proteins and DNA structures have been reported to form
synthetic ion channels.[19] Since only a few guanosine derivatives
have been evaluated for transporting metal ions across the
biological membrane, we have studied the ability of lipophilic 8-
bromoguanosine, G1 to form ion channels. The voltage-clamp
experiments were carried out on planar solvent-free bilayers in 1
M KCl, phosphate (2 mM) buffer at pH 7.4. First, giant
unilamellar vesicles (GUVs) were prepared from 1,2-
Figure 7. Self-assembly of guanosine derivatives to form ion-channels in
biological membrane.
In summary, we have demonstrated that guanosine
derivative G1 self-assemble to form distinct nanoarchitectures.
The lipophilic TBDMS protected bromoguanosine derivative
forms different nanostructures with time, exhibits birefringence in
its crystal state and binds dye molecules. It can also transport
ions through the membrane. This study reveals that simple
monoguanosine derivatives can be used to build functional
nanoarchitectures as well as membrane spanning channels for
ion transportation.
diphytanoyl-sn-glycero-3-phosphocholine
(DPhPC)
and
cholesterol (9:1). Then planar solvent-free bilayers were
[20]
generated by spreading GUVs on a small aperture in glass.
Guanosine derivative G1 (20 µM) was added to the cis-side of
the chamber and current traces were recorded at an applied
voltage of -100 mV.
ACKNOWLEDGMENTS
We thank Department of Atomic Energy (BNRS) and DST India for
research funding. RND thanks CSIR India and DAAD exchange
programme for research fellowships. JD thanks DST for
SwarnaJayanti fellowship.
a
Keywords: • biomimetic ion channel • birefringence • dye
binding • lipophilic guanosine • self-assembly• phospholipid
bilayer
Figure 6. Traces of single-channel recordings obtained by the insertion of (a)
G1 (20 µM) to the cis side of the chamber after the planar bilayer was formed.
The experiment was performed at -100 mV in 1 M KCl, phosphate (2 mM)
buffer at pH 7.4. Statistical distribution of the observed conductance states for
[1]
a) J. T. Davis, Angew. Chem. Int. Ed. 2004, 43, 668-698; b) K. Araki, I.
Yoshikawa, Top. Curr. Chem. 2005, 256, 133-165; c) S. Sivakova, S. J.
Rowan, Chem. Soc. Rev. 2005, 34, 9-21; d) J. T. Davis, G. P. Spada,
Chem. Soc. Rev. 2007, 36, 296-313; e) S. Lena, S. Masiero, S.
Pieraccini, G. P. Spada, Chem. Eur. J. 2009,15, 7792-7806; f) J. Dash,
P. Saha, Org. Biomol. Chem. 2016, 14, 2157-2163; g) G. M. Peters, J.
T. Davis, Chem. Soc. Rev. 2016, 45, 3188-3206.
(b) G1 (592 events).
The characteristic current steps were observed upon
insertion of G1 into the lipid membrane. These results suggest
[
2]
a) G. Gottarelli, S. Masiero, E. Mezzina, S. Pieraccini, J. P. Rabe, P.
Samorí, G. P. Spada, Chem. Eur. J. 2000, 6, 3242–3248; b) K. Araki, R.
Takasawa, I. Yoshikawa, Chem. Commun. 2001, 1826-1827; c) T.
Giorgi, F. Grepioni, I. Manet, P. Mariani, S. Masiero, E. Mezzina, S.
Pieraccini, L. Saturni, G. P. Spada, G. Gottarelli, Chem. Eur. J. 2002, 8,
the formation of distinct channels (Figure 6a, Table S1) that can
+
transport K ions across the bilayer. The opening and closing of
these ion channels indicate a dynamic self-association process
of guanosine derivatives. The histogram analysis of the current
2143-2152; d) S. Lena, M. A. Cremonini, F. Federiconi, G. Gottarelli, C.
Graziano, L. Laghi, P. Mariani, S. Masiero, S. Pieraccini, G. P. Spada,
Chem. Eur. J. 2007, 13, 3441-3449; e) M. El Garah, R. C. Perone, A. S.
Bonilla, S. Haar, M. Campitiello, R. Gutierrez, G. Cuniberti, S. Masiero,
A. Ciesielski, P. Samorì, Chem. Commun. 2015, 51, 11677-11680.
a) G. W. Collie, G. N. Parkinson, Chem. Soc. Rev. 2011, 40, 5867-
traces showed that G1 formed pores with
a maximum
conductance value of 1.2 nS (Figure 6b) and it also frequently
formed channels with conductance values of 0.1-0.5 nS (60.1 %)
[3]
(
Table S1). Owing to weak lipophilicity and low solubility, simple
5
892; b) A. Bugaut, S. Balasubramanian, Nucleic Acids Res. 2012, 40,
4727-4741; c) P. Murat, S. Balasubramanian, Curr. Opin. Genet. Dev.
014, 25, 22-29; d) S. Neidle, J. Med. Chem. 2016, 59, 5987-6011.
guanosine 1 and 8-bromoguanosine 2 (Figure 1) failed to form
ion channels, whereas in case of G1, the lipophilic TBDMS
groups enabled the insertion of G1 nanostructures into the lipid
bilayer resulting in significant conductance values. The most
frequently observed channels with conductance values (0.1-0.5
2
[
4]
5]
a) J. L. Sessler, M. Sathiosatham, K. Doerr, V. Lynch and K. A. Abboud,
Angew. Chem., Int. Ed. 2000, 39, 1300–1303; b) L. E. Buerkle, H. A.
von Recum, S. J. Rowan, Chem. Sci. 2012, 3, 564-572.
R. N. Das, Y. P. Kumar, S. Pagoti, A. J. Patil, J. Dash, Chem. Eur. J.
2012, 18, 6008-6014
[
nS) indicate the formation of G-quartet like ion channels in the
lipid bilayer (Figure 7).[15a,c] The CD spectrum of G1, embedded
[6]
7]
Y. -F. Gao, Y. -J. Huang, S. -Y. Xu, W. -J. Ouyang, Y. -B. Jiang,
Langmuir 2011, 27, 2958-2964.
in the membrane showed a positive peak at 253 nm and
negative peak at 275 nm, suggesting the formation of G-quartet
like structures in the membrane (Figure S3, S.I.). The G-quartets
could stack into columns in the lipid bilayer enabling transport of
[
CCDC 1582231 contains the supplementary crystallographic data of G1.
The data can be obtained free of charge from the Cambridge
Crystallographic Data Centre via www.ccdc.cam.ac.uk/getstructures.
a) G. Reddy, A. Marsh, J. T. Davis, S. Masiero, S. P. Brown,
Cryst. Growth Des. 2015, 15, 5945-5954; b) A. L. -Lior, B. Pokroy, B. L.
[8]
+
K ions through the G-quartet pores (Figure 7).
-Sivan,
L.
Leiserowitz,
S.
Weiner,
L.
Addadi,
Cryst. Growth Des. 2008, 8, 507-511; c) S. N. Fejer, D. Chakrabartia, D.
This article is protected by copyright. All rights reserved.

Chemistry - A European Journal
10.1002/chem.201800205
COMMUNICATION
J. Wales, Soft Matter 2011, 7, 3553-3564; d) D. Gur, B. A. Palmer, S.
Weiner, L. Addadi, Adv. Funct. Mater. 2017, 27, 1603514.
rd
[
9]
a) E. Hecht, Optics, 3 ed., Addison-Wesley, New York, 1998; b) S.
Tadepalli, J. M. Slocik, M. K. Gupta, R. R. Naik, S. Singamaneni, Chem.
Rev., 2017, 117, 12705-12763; c) M. K. -Tani, T. Tani, S. B. Mehta, A.
Verma, R. Oldenbourg, Mol. Reprod. Dev. 2015, 82, 548-562.
[
[
10] M. S. Kaucher, Y. -F. Lam, S. Pieraccini, G. Gottarelli, J. T. Davis,
Chem. Eur. J. 2005, 11, 164-173.
11] a) G. M. Peters, L. P. Skala, J. T. Davis, J. Am. Chem. Soc. 2016, 138,
134-139; b) J. Dash, A. J. Patil, R. N. Das, F. L. Dowdall, S. Mann, Soft
Matter 2011, 7, 8120–8126.
[
[
[
12] M. S. Kaucher, W. A. Harrell, Jr., J. T. Davis, J. Am. Chem. Soc. 2006,
128, 38.
13] N. Sakai, Y. Kamikawa, M. Nishii, T. Matsuoka, T. Kato, S. Matile, J.
Am. Chem. Soc. 2006, 128, 2218-2219.
14] a) L. Ma, M. Melegari, M. Colombini, J. T. Davis, J. Am. Chem. Soc.
2
008, 130, 2938-2939; b) L. Ma, W. A. H. Jr, J. T. Davis, Org. Lett.
009, 11, 1599-1602
2
[
15] a) Y. P. Kumar, R. N. Das, S. Kumar, O. M. Schütte, C. Steinem, J.
Dash, Chem. Eur. J. 2014, 20, 3023-3028; b) R. N. Das, Y. P. Kumar,
O. M. Schꢀtte, C. Steinem, J. Dash, J. Am. Chem. Soc. 2015, 137, 34-
37; c) Y. P. Kumar, R. N. Das, O. M. Schütte, C. Steinem, J. Dash,
Nature protocols 2016, 11, 1039-1056.
[16] a) G. W. Gokel, A. Mukhopadhyay, Chem. Soc. Rev. 2001, 30, 274-
2
8
86; b) N. Sakai, J. Mareda, S. Matile, Acc. Chem. Res. 2005, 38, 79-
7; c) G. W. Gokel, I. A. Carasel, Chem. Soc. Rev. 2007, 36, 378-389;
d) T. Saha, S. Dasari, D. Tewari, A. Prathap, K. M. Sureshan, A. K.
Bera, A. Mukherjee, P. Talukdar, J. Am. Chem. Soc. 2014, 136, 14128-
14135; e) S. -K. Ko, S. K. Kim, A. Share, V. M. Lynch, J. Park, W.
Namkung, W. Van Rossom, N. Busschaert, P. A. Gale, J. L. Sessler, I.
Shin, Nat. Chem. 2014, 6, 885-892; f) J. R. Burns, A. Seifert, N. Fertig,
S. Howorka, Nat. Nanotechnol. 2016, 11, 152-156; g) N. Busschaert, S.
-H. Park, K. -H. Baek, Y. P. Choi, J. Park, E. N. W. Howe, J. R. Hiscock,
L. E. Karagiannidis, I. Marques, V. Félix, W. Namkung, J. L. Sessler, P.
A. Gale, I. Shin, Nat. Chem. 2017, 9, 667-675; g) S. Howorka, Nat.
Nanotechnol. 2017, 12, 619-630.
[
[
[
17] a) C. J. Drummond, C. Fong, Curr. Opin. Colloid Interface Sci. 2000, 4,
449-456; b) L. Leanza, A. Managò, M. Zoratti, E. Gulbins, I. Szabo,
Biochim. Biophys. Acta. 2016, 1863, 1385-1397.
18] a) Y. Choi, L. A. Baker, H. Hillebrenner, C. R. Martin, Phys. Chem.
Chem. Phys. 2006, 8, 4976-4988; b) C. R. Martin, Z. S. Siwy, Science
2007, 317, 331-332.
19] a) P. H. Schlesinger, R. Ferdani, J. Liu, J. Pajewska, R. Pajewski, M.
Saito, H. Shabany, G. W. Gokel, J. Am. Chem. Soc. 2002, 124, 1848-
1849; b) J. S. -Quesada, M. P. Isler, M. R. Ghadiri, J. Am. Chem. Soc.
2002, 124, 10004-10005; c) H. Itoh, S. Matsuoka, M. Kreir, M. Inoue, J.
Am. Chem. Soc. 2012, 134, 14011-14018; d) M. Langecker, V. Arnaut,
T. G. Martin, J. List, S. Renner, M. Mayer, H. Dietz, F. C. Simmel,
Science 2012, 338, 932-936; e) N. Sakai, S. Matile, Langmuir 2013, 29,
9031-9040; f) M. Langecker, V. Arnaut, J. List, F. C. Simmel, Acc.
Chem. Res. 2014, 47, 1807-1815; g) T. Saha, A. Gautam, A.
Mukherjee, M. Lahiri, P. Talukdar, J. Am. Chem. Soc. 2016, 138,
1
6443-16451; h) B. P. Benke, P. Aich, Y. Kim, K. L. Kim, M. R. Rohman,
S. Hong, I. -C. Hwang, E. H. Lee, J. H. Roh, K. Kim, J. Am. Chem. Soc.
017, 139, 7432-7435;
2
[
20] a) A. Kurz, A. Bunge, A. -K. Windeck, M. Rost, W. Flasche, A.
Arbuzova, D. Strohbach, S. Müller, J. Liebscher, D. Huster, A.
Herrmann, Angew. Chem. Int. Ed. 2006, 45, 4440-4444; b) L. Mathivet,
S. Cribier, P. F Devaux, Biophys. J. 1996, 70, 1112-1121; c) M. I.
Angelova, D. S. Dimitrov, Faraday Discuss. Chem. Soc. 1986, 81, 303-
311.
This article is protected by copyright. All rights reserved.

Chemistry - A European Journal
10.1002/chem.201800205
COMMUNICATION
Entry for the Table of Contents
Layout 1:
COMMUNICATION
[
a]
[a]
Rabindra Nath Das, Y. Pavan Kumar,
S. Arun Kumar,[ Ole Mathis Schütte,
Claudia Steinem,[b] Jyotirmayee Dash[a]*
a]
[b]
Page No. – Page No.
Self-assembly of a Guanosine
Derivative to Form Nanostructures
and Transmembrane Channels
Nucleoside nanostructure and ion channel: A simple lipophilic nucleoside
analogue forms self-assembled structures in solution, solid state and in lipid bilayer.
Interestingly, the self-assembled structure of this nucleoside exhibits birefringence,
incorporates a bioactive dye and forms transmembrane ion channels across a
phospholipid bilayer.
This article is protected by copyright. All rights reserved.