Smart hydrogels from laterally-grafted peptide assembly

Article abstract of DOI:10.1039/c2cc34528e

Small peptides carrying laterally-grafted azobenzene units self-assemble into photo-responsive hydrogels which are applied as a smart matrix for controlling the dye molecules release. We demonstrate that a delicate balance among peptides interactions plays a pivotal role in the photo-responsive gel-sol transition.

Full text of DOI:10.1039/c2cc34528e

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Cite this: Chem. Commun., 2012, 48, 8796–8798
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COMMUNICATION
Smart hydrogels from laterally-grafted peptide assemblyw
Wen Li,ab Il-soo Park,a Seong-Kyun Kanga and Myongsoo Lee*a
Received 25th June 2012, Accepted 13th July 2012
DOI: 10.1039/c2cc34528e
Small peptides carrying laterally-grafted azobenzene units self-
assemble into photo-responsive hydrogels which are applied as a
smart matrix for controlling the dye molecules release. We
demonstrate that a delicate balance among peptides interactions
plays a pivotal role in the photo-responsive gel–sol transition.
sodium hydroxide21 or enzyme.22 The main reason is that the
intermolecular interactions among b-sheet peptides are too
strong to be broken by the conformational transition of the
azobenzene units. This character led us to consider that
weakening the interactions by optimizing the molecular design
should be a rational strategy for creating very simple photo-
responsive hydrogels. With this in mind, we designed two
small peptides (1 and 3) containing light sensitive azobenzene
units in a lateral way and terminal lysine groups (Fig. 1). It is
expected that the electrostatic repulsions between protonated
lysine groups reduce the secondary interactions of peptides
and result in simple photo-responsive gels. Peptide 2 with
glutamic acid at one terminal was synthesized for elucidating
the importance of the electrostatic repulsions. Herein, we
present the photo-responsive self-assembly and the gelation
behavior of the synthesized peptides. We also demonstrated
that the smart hydrogel can be used further as a vehicle to
control the dye molecules release.
Supramolecular hydrogels with a stimuli responsive functionality
have been of immense interest to the field of nano-scale materials.1,2
This interest stems from the fact that the responsive gels allow
the development of smart biomaterials, such as controlled
drug delivery, sensors, and tissue engineering.3,4 Smart hydrogels
obtained from natural biomolecules are appealing since
they are more likely to be biodegradable, biocompatible, and
bioresorbable.5,6 Rational peptide design and engineering are
thus emerging as particularly promising routes to such materials
due to the relative easiness in their synthesis.7–9 An essential
requirement is that the peptides should have a propensity to form
1D self-assembly nanofibers, which can entangle to form 3D
networks with high water content.10 To advance the utility of
smart hydrogels, a lot of effort has been focused on triggering the
self-assembly and gelation behavior of small peptides in various
stimuli-responsive manners, such as pH,11 temperature,12 ionic
strength,13 oxidation–reduction,14 enzymatic manipulation,15
and light.16 Among them, light, especially the photoisomerization
of azobenzene, is an interesting choice as it can be manipulated
spatially and temporally in a non-contact fashion,17,18 and the
photochromic reaction is repeatable at will without apparent
fatigue.19
Small peptides with b-sheet structures are commonly favourable
for the formation of 1D nanofibers due to the directional hydrogen
bonding within the peptide backbones.20 Although a few b-sheet
peptides with azobenzene units have been synthesized to create
smart hydrogels, the photo-responsive gel–sol transitions are
complicated.21,22 In other words, UV-light irradiation is not
enough to destroy the peptide 3D networks, and the gel to sol
transition process needs additional assistance reagents, such as
a Center for Bio-Responsive Assembly, Department of Chemistry,
Seoul National University, Seoul 151-747, Korea.
E-mail: myongslee@snu.ac.kr; Fax: +82 2-393-6096;
Tel: +82 2-880-4340
b State Key Laboratory of Supramolecular Structure and Materials,
Jilin University, Changchun 130012, China
w Electronic supplementary information (ESI) available: Experimental
details of peptides synthesis and characterizations, CD, FT-IR and
UV spectra, additional TEM and SEM images. See DOI: 10.1039/
c2cc34528e
Fig. 1 Molecular structures of the synthesized peptides and the
schematic presentation of the photo-responsive self-assembly structures
of peptide 1.
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conformational change enforces long nanofibers to break up into
discrete spherical aggregates (Fig. 2c and d). Consequently, the
disassociation of the nanofibers caused by photoisomerization of
azobenzene units drove the gels to transform into a fluid solution
(Fig. 2d, inset). After aging the solution of spherical structures for
two hours under daylight, long nanofibers and self-supporting
hydrogels were observed again (Fig. S5, ESIw), indicating
a reversible sol–gel transition with light. Interestingly, circular
dichroism (CD) spectra showed that the b-sheet structure of
peptide 1 remained almost unchanged after UV irradiation
(Fig. 3a), indicating that the photoisomerization of azobenzene
did not cause a significant variation in the backbone of 1. It is
clear that the stacking of azobenzene groups has an important
influence on the self-assembly structures even though the
backbone structure of peptide 1 remains constant. Actually,
the hydrophobic azobenzene moieties have a strong tendency
to associate with each other to minimize their contact with
water. The planar geometry of trans-azobenzene side groups
provides an energetic contribution as well as the directional
hydrogen bonding for the formation of crossed b-sheet structures,
which further stack into extended 1D structure. When exposing
the solution of nanofibers to UV irradiation, the nonplanar cis
azobenzene units caused an increased bulky effect, which does not
facilitate the peptides alignment into 1D arrangement. This result
is similar to the self-assembly of aromatic dipeptides reported
by Reches and Gazit.25 Combining with the electrostatic
repulsions between protonated lysine groups, these factors
resulted in random b-sheet structures, which were not competent
for 1D growth. Some molecular simulations suggested that the
random b-sheets had a compromise propensity to adopt a spherical
interfacial curvature.26,27 We also studied the photo-responsive
gelation behavior of 1 after treatment with salt and basic reagents.
In those cases, the electrostatic repulsions between protonated
lysine groups would be suppressed significantly. The cgc value
of peptide 1 is down to 3.4 wt% and 3.1 wt% for ammonia
water (pH B 10.5) and salt solution (10 mM NaCl), respectively.
However, the resulting gels did not transform into fluid solution
even after exposing to UV light for 30 min (Fig. S6, ESIw),
implying that the enhanced peptides interactions are strong
enough to inhibit the disassociation of the nanofibers.
Fig. 2 Photo-responsive self-assembly structures of peptide 1:
(a) TEM image of 1 in 40 mM aqueous solution; (b) SEM image of
1 gel (inset, optical image of 1 gel); (c) TEM image of 1 solution and
(d) SEM image of 1 gel sample (inset, optical image of 1 sol) after
exposing to 360 nm UV light for 15 min.
A transmission electron microscopy (TEM) image revealed
that peptide 1 self-assembled into long nanofibers with a
uniform diameter of 3 nm (Fig. 2a and inset) in diluted
aqueous solution. With increasing concentration, the nanofibers
of peptide 1 entangled with each other to form 3D networks
(Fig. 2b), corresponding to stable hydrogel formation (Fig. 2b,
inset). The critical gel concentration (cgc) of 1 is 5.1 wt% based
on the inverting tube method. The high cgc value is probably
due to the presence of electrostatic repulsions between the
protonated lysine groups (pKa B 10.5) in aqueous solution,
which depresses the aggregation to some extent.23
Circular dichroism (CD) spectra of peptide 1 (Fig. 3a)
displayed a negative Cotton effect at 220 nm, indicative of
typical b-sheet conformation. Fourier transform infrared (FT-IR)
spectra of the gels (Fig. 3b) showed well-defined amide I bands
centered at 1629 cmÀ1 and 1679 cmÀ1, which suggests that the
nanofibers of peptide 1 consist of antiparallel b-strands.24 The
X-ray diffraction of the peptide 1 xerogels (Fig. S3, ESIw)
indicated layered structures with a distance (d) of 3.03 nm, which
is consistent with the TEM observation. The molecular length (l)
between the hydrophilic amine at the e-position of lysine and
the azobenzene units was estimated to be 2.0 nm. Considering
the obtained layer spacing located at l o d o 2l, we propose
that the nanofibers consist of a bilayer structure with inter-
calated azobenzene units, as shown in Fig. 1.
To gain further insight into the strength of b-sheets of
peptide 1, we have prepared the analogous one bearing a
glutamic acid at the one end of the molecules (Fig. 1, peptide 2).
This molecular design would lead to enhanced b-sheet strength
through increased electrostatic interactions between glutamic
acid and lysine units of the adjacent molecules (Fig. S7, ESIw).
Indeed, peptide 2 self-assembles into rigid nanofibers (Fig. 4a),
which further form 3D networks and stable gels (Fig. 4b and
inset) at lower concentrations (B2.6 wt%). However, the gels
do not show responsive characteristics after transformation of
the azobenzene units into cis-conformation by UV irradiation
(Fig. S8, ESIw). The TEM (Fig. 4c), SEM (Fig. 4d), and
optical images (Fig. 4d, inset) show that the fibrillar structures,
3D network structures, and the gel matrix remain unchanged,
respectively, even after UV irradiation for long time (above
30 min).
Notably, the gels of 1 are responsive to UV irradiation
through a conformational transition of azobenzene moieties
from trans to cis states (see UV spectra, Fig. S4 (ESIw)). This
The above results strongly demonstrate that construction
of photo-responsive hydrogels requires a delicate balance
among peptide interactions. In our system, the introduction
Fig. 3 (a) Circular dichroism (CD) spectra of peptide 1 aqueous
solution before (black line) and after (red line) UV irradiation;
(b) FT-IR spectrum of peptide 1 gel.
c
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not show responsive characteristics any more even after UV
irradiation for longer time. The results here suggest that
construction of photo-responsive hydrogels requires appropriate
secondary interactions of peptides. The photo-responsive
characteristics of such peptide gels allowed us to further tune
the release of dye molecules with UV light. Although the
present smart gels still need to be improved in the future
for applying under physiological conditions, we believe that
this report will provide an important insight into rational
molecular design for photo-responsive peptide hydrogels.
We gratefully acknowledge the National Research Foundation
of Korea (NRF) grant funded by the Korean government
(MEST) (No. 2012-0001240 and 2011-35B-C00024).
Fig. 4 Self-assembly structures of peptide 2: (a) TEM image of 2 in
40 mM aqueous solution; (b) SEM image of 2 gel (inset, optical image
of 2 gel); (c) TEM image of 2 solution and (d) SEM image of 2 gel
sample (inset, optical image of 2 gel) after exposing to 360 nm UV light
for 30 min.
Notes and references
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Fig. 5 Time course of the rhodamine B release, with (cycles) and
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of electrostatic repulsions into b-sheet peptides is a simple
strategy to design smart hydrogels. To further support this
proposal, we have synthesized another peptide with more
azobenzene units (3). Peptide 3 formed stable gels above the
concentration of 4.03 wt%, which showed responsive char-
acteristics after transformation of the azobenzene units into
cis-conformation by UV irradiation (Fig. S9, ESIw).
With the photo-responsive characteristics, the gels are
considered as a smart material to control the dye release.
Rhodamine B (RhB) was firstly mixed with the peptide 1 gel.
We then investigated the release of RhB to distilled water with
and without UV irradiation by detecting the emission intensity
of RhB at 581 nm. As shown in Fig. 5, the emission intensity
within 10 hours is near 125 in the absence of UV irradiation.
In contrast, when applying the UV irradiation, a more rapid
release of RhB is observed in the composite hydrogels together
with the moderate decrease in the gel volume, because of the
gel-to-sol phase transition, causing the gradual release of RhB
molecules embedded in the gel matrix.
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with laterally-grafted azobenzene units. Peptides 1 and 3 with
lysine groups at both terminals formed photo-responsive
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repulsions between protonated lysine groups. When enhancing
the b-sheet strength through the introduction of ammonia water,
salts, or electrostatic interaction, the resulting hydrogels did
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c
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