A light-regulated synthetic ion channel constructed by an azobenzene modified hydraphile

Article abstract of DOI:10.1016/j.cclet.2015.05.010

Biological ion channels are key molecules for cellular regulation and communication. To mimic the structure and functions of nature ion channels, a new class of light-regulated transmembrane ion channels was reported based on tri(macrocycle) hydraphile an

Full text of DOI:10.1016/j.cclet.2015.05.010

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CCLET 3314 1–6
Chinese Chemical Letters xxx (2015) xxx–xxx
Contents lists available at ScienceDirect
Chinese Chemical Letters
journal homepage: www.elsevier.com/locate/cclet
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Original article
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A light-regulated synthetic ion channel constructed by an azobenzene
modified hydraphile
* *
Q1 Rong-Yan Yang, Chun-Yan Bao , Qiu-Ning Lin, Lin-Yong Zhu
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Shanghai Key Laboratory for Functional Materials Chemistry, Institute of Fine Chemicals, East China University of Science and Technology, Shanghai 200237,
China
A R T I C L E I N F O
A B S T R A C T
Article history:
Biological ion channels are key molecules for cellular regulation and communication. To mimic the
structure and functions of nature ion channels, a new class of light-regulated transmembrane ion
channels was reported based on tri(macrocycle) hydraphile and azobenzene photoswitch (hydraphile 1).
The liposome-based proton transport assays showed that hydraphile 1 exhibited excellent transmem-
Received 9 March 2015
Received in revised form 1 April 2015
Accepted 9 April 2015
Available online xxx
brane activity (Y), and Ymax arrived 0.7 at 40
mmol/L. The successful isomerization of azobenzene moiety
was confirmed and qualified by UV and NMR spectra. Upon alternative irradiation of 365 nm UV light
and 450 nm visible light, the transmembrane activity of hydraphile 1 was regulated between 0.35 and
0.5, reversibly. All the obtained results have demonstrated the promise of developing excellent synthetic
ion channels with ion gating properties based on simple molecular design.
Keywords:
Synthetic ion channel
Light-regulated ion transport
Hydraphiles
ß 2015 Chinese Chemical Society and Institute of Materia Medica, Chinese Academy of Medical Sciences.
Published by Elsevier B.V. All rights reserved.
Azobenzene photoisomerization
Self-assembly
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1. Introduction
polarity changes. However, this approach requires complex 29
fabrication steps, involves design difficulties, and is sometimes 30
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Amongst various forms of input stimuli, light stands out as one
of the most promising stimulation modalities for use in life science
research because it offers nonphysical contact, controllable
intensity, and high temporal and spatial precision. Recently, the
use of light has been greatly improved in bioimaging, photo-
regulation of gene expression, photo-release of drug, and light-
gated ion channel transport [1,2]. Although light-sensitive ion
translocation (rhodopsin [3]) is limited in natural occurring
organisms, light-gated ion channels can enable the direct
manipulation of cellular excitability in genetically modified cells
because cell activation can be directly linked to the diffusion of
ions across the cell membrane [4].
Over the last few decades, many light-regulated artificial ion
channels have been reported [5–9]. There are now two principal
strategies for constructing light-responsive ion channels. The first
strategy uses ‘‘optochemical genetics’’ [5,6], where the channel
protein is genetically encoded with a photoreceptor that under-
goes a photochemical reaction, moving in or out of the ion channel
(ON or OFF) to gate ion flow by way of major conformation or
unpredictable. Thus, it can be used on only a few well- 31
characterized protein channels. The alternative approach uses 32
synthetic light-responsive ion channels by attaching a photoswitch 33
to the channel molecules, which then regulates ion translocation 34
by reversibly changing the channel structures or ion dipole 35
interactions [7–9]. Azobenzene is one of the most commonly used 36
photoswitches due to a large change in azobenzene length and 37
geometry upon photoisomerization from a linear and mostly 38
planar trans form to a kinked 3D cis form, in addition to its short 39
response time [10]. Several successful light-regulated ion channels 40
were constructed due to the reversible photoregulation of 41
azobenzene [7a,11].
42
‘‘Hydraphiles’’, so called because they are amphiphiles and they 43
are two-headed and reminiscent of the mythical hydra serpent, 44
represent one kind of typical single-molecular channel models 45
devoted by Gokel et al. [12]. The hydraphiles have three diaza-18- 46
crown-6 residues linked and terminated by alkyl chains, in which 47
the inner macroring embeds in the membrane and the two 48
terminal rings are near the bilayer surface by supramolecular 49
interactions between lipid and hydraphile. Research shows that 50
the terminal substitution and the spacer length have great effect on 51
the transport activity [12e,f], and the closer of the spacer lengths 52
are to the thick of bilayer lipid membrane, the more efficient of the 53
transport activity of the hydraphiles. Meanwhile, several of the 54
*
Corresponding authors.
Q2
E-mail addresses: baochunyan@ecust.edu.cn (C.-Y. Bao),
linyongzhu@ecust.edu.cn (L.-Y. Zhu).
http://dx.doi.org/10.1016/j.cclet.2015.05.010
1001-8417/ß 2015 Chinese Chemical Society and Institute of Materia Medica, Chinese Academy of Medical Sciences. Published by Elsevier B.V. All rights reserved.
Please cite this article in press as: R.-Y. Yang, et al., A light-regulated synthetic ion channel constructed by an azobenzene modified
hydraphile, Chin. Chem. Lett. (2015), http://dx.doi.org/10.1016/j.cclet.2015.05.010

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R.-Y. Yang et al. / Chinese Chemical Letters xxx (2015) xxx–xxx
Scheme 1. The schematic presentation of the structures and transmembrane ion transport of hydraphile 1.
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hydraphile channels are biologically active and show antibacterial
[12g], and anticancer toxic behavior [12a,h]. Taking inspiration
from the work of Gokel et al., we used the classic backbone of the
‘‘hydraphiles’’ as the transmembrane channel and introduced
azobenzene as the photoswitch to construct light-regulated ion
channel (hydraphile 1). As shown in Scheme 1, the photoisome-
rization of azobenzene induced the change of the transmembrane
length of the channel, thus regulated the ion transport reversibly.
assay were as same as the description in our previous report
[11a,b]. Here, the mixture of EYPC and cholesterol (10:1, w:w) was
used for the membrane of LUVs, and 8-hydroxy-1,3,6-pyrenetri-
sulfonate (HPTS) was used as the pH-sensitive fluorescent probe,
the final concentration of the lipids in the experiments was
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liposomes), and the size of the vesicles was around 150 nm. In
the time-dependent change in fluorescence intensity, 30 L,
0.5 mol/L KOH was added at t = 50 s, 30 L transporter in DMSO
with different concentrations was added at t = 100 s, and 60 L of
m
mol/L (assuming 100% of lipids were incorporated into
m
m
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2. Experimental
m
5% Triton X-100 aqueous solution was added at t = 350 s for final
completed balance. Time courses of fluorescence intensities It were
obtained by first, ratiometric analysis (R = It,450/It,405) and second,
normalization according to Eq. (1),
2.1. Materials
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All starting materials were obtained from commercial suppliers
and were used without further purification unless otherwise
stated. All air- or moisture-sensitive reactions were performed
using oven-dried or flame-dried glassware under an inert
atmosphere of dry argon. Egg yolk phosphatidylcholine (EYPC)
was obtained from Avanti Polar lipids as a solution in chloroform
(25 mg/mL).
ðR ꢀ R100Þ
It ¼
(1)
ðR1 ꢀ R100Þ
where R100 = R before addition of transporter and R = R after
1045
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1
addition of Triton X-100. It at 350 s just before addition of Triton X-
100 was defined as transmembrane activity Y.
2.4. Synthesis of hydraphile 1
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2.2. Characterizations
Compound 8: Under Ar gas protection, compound 9 (0.2 g,
0.76 mmol) was dissolved in 10 mL dry CH2Cl2. Sodium hydride
(73 mg, 3.05 mmol) was added slowly and stirred for 15 min at
room temperature, then bromoacetyl bromide (0.17 mL in 7 mL
dry CH2Cl2) was added dropwise to the flask and stirred for another
6 h at room temperature (Scheme 2). The reaction solution was
concentrated in vacuum and chromatographed on a column of
silica (silica gel, 20% methanol/CH2Cl2) to obtain 0.28 g (73%)
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Proton and carbon nuclear magnetic resonance spectra (1H
NMR, 13C NMR) were recorded on a Bruker Avance 500 (400 MHz)
spectrometer. Mass spectra were recorded on a Micromass GCTTM
and
a Micromass LCTTM. Fluorescence measurements were
performed on a Varian Cary Eclipses fluorescence spectrometer
equipped with a stirrer and a temperature controller (kept at 25 8C
unless otherwise noted). Absorption spectra were recorded on a
Shimadzu UV-2550 UV-vis spectrometer. A Mini-Extruder used for
the preparation of large unilamellar vesicles (LUVs) was purchased
from Avanti Polar lipids. The size of EYPC vesicles was determined
using a DelsaTM Nano Submicron Particle Size and Zeta Potential
Particle Analyzer (Beckman Coulter Inc., USA). A 365 nm LED lamp
(30 mW/cm2) was used for photolysis of compounds and photo-
controlled experiments.
compound 8 as a colorless liquid. 1H NMR (400 MHz, CDCl3):
(d, 4H, J = 3.1 Hz), 3.75 (d, 4H, J = 5.3 Hz), 3.71–3.56 (m, 20H). 13C
NMR (100 MHz, CDCl3): 170.44, 168.71, 70.95, 70.81, 70.70,
d 3.96
d
70.45, 69.82, 69.54, 69.17, 69.10, 50.53, 50.50, 48.54. MS (EI): m/z
Calcd. for C16H28Br2N2O6 [M]+: 504.2. Found: 504.2.
Compounds
references [11c,a]. Compound 7: 1H NMR (400 MHz, CDCl3):
7.34 (s, 5H), 3.91 (s, 4H), 3.63 (d, 6H, J = 14.4 Hz), 3.52 (d, 8H,
J = 14.4 Hz), 3.14 (s, 4H), 2.73 (s, 4H). 13C NMR (CDCl3, 100 MHz):
7 and 5 were synthesized as the reported
d
87
2.3. HPTS assay
d
138.47, 128.98, 128.32, 127.04, 70.09, 69.91, 68.36, 66.69, 57.04,
56.00, 48.82. MS (EI): m/z Calcd. For C19H32N2O4 [M]+:
88
89
Preparation of large unilamellar vesicles (LUVs) and determi-
nation the transport activity of the compounds with the HPTS
Please cite this article in press as: R.-Y. Yang, et al., A light-regulated synthetic ion channel constructed by an azobenzene modified
hydraphile, Chin. Chem. Lett. (2015), http://dx.doi.org/10.1016/j.cclet.2015.05.010

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Scheme 2. Synthesis of hydraphile 1. Reagents and conditions: (a) bromoacetyl bromide/NaH, CH2Cl2, 73% yield; (b) benezyl bromide, K2CO3, 46% yield; (c) H2O/KOH, 64%
yield; (d) tert-butyl bromoacetate, K2CO3, acetonitrile, 56% yield; (e) 8, K2CO3/TBAB, acetonitrile, 93% yield; (f) TFA/CH2Cl2, 96% yield; (g) 1-chloro-N,N,2-
trimethylpropenylamine, CH2Cl2; 7, TEA/CH2Cl2, 68% yield.
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352.2. Found: 352.2. Compound 5: 1H NMR (400 MHz, DMSO-d6):
10.13 (s, 2H), 7.87–7.57 (m, 4H), 6.95–6.87 (m, 4H). 13C NMR
d
114.90, 82.69, 70.21, 68.42, 64.73, 41.02. MS (ESI): m/z Calcd. for 166
44H50N6O14 [M+H]+: 887.3385. Found: 887.3385.
167
Hydraphile 1: Under Ar gas protection, compound 2 (0.2 g, 168
C
(100 MHz, DMSO-d6): d 165.22, 150.49, 129.40, 121.02. MS (EI): m/
z Calcd. for C12H10N2O2 [M]+: 214.0742. Found: 214.0741.
0.23 mmol) was dissolved in 10 mL dry CH2Cl2. Then 0.5 mL 1- 169
chloro-N,N,2-trimethylpropenylamine was added, and stirred for 170
3 h at room temperature. After evaporation solvent, the interme- 171
diate product (0.17 g) was added to next step directly. Under Ar gas 172
protection, intermediate product (0.2 g, 0.57 mmol) and triethy- 173
lamine were dissolved in 8 mL dry CH2Cl2. Then a solution of 174
compound 7 in 8 mL dry CH2Cl2 was added dropwise to this 175
reaction system, and stirred for 8 h at room temperature. After 176
removing solvent, the product was chromatographed on a column 177
of silica (silica gel, 2% methanol/CH2Cl2) to obtain 0.24 g (68%) 178
Compound 4: Compound 5 (1 g, 4.67 mmol) and potassium
carbonate (1.9 g, 14 mmol) were dissolved in dry acetonitrile. A
solution of tert-butyl bromoacetate (0.69 g, 4.67 mmol) in 10 mL
dry acetonitrile was added dropwise to the reaction system. The
reaction kept refluxing for 10 h. After cooling to room temperature,
and then filtered. After evaporation solvent, the product was
chromatographed on a column of silica (silica gel, 0.5% methanol/
CH2Cl2) to obtain 0.85 g (56%) compound 4 as an orange powder.
1H NMR (400 MHz, CDCl3):
6.89 (m, 4H), 4.60 (s, 2H), 1.50 (s, 9H). 13C NMR (101 MHz, CDCl3):
d 7.87 (dt, 4H, J = 13.1, 5.9 Hz), 7.04–
d
compound as an orange powder. 1H NMR (400 MHz, CDCl3):
(d, 8H, J = 8.1 Hz), 7.31 (s, 10H), 7.04 (d, 8H, J = 8.1 Hz), 4.85 (s, 8H), 180
3.78–3.59 (m, 76H). 13C NMR (100 MHz, CDCl3):
168.02, 165.23, 181
d 7.85 179
167.99, 159.65, 158.21, 147.51, 146.98, 124.65, 124.32, 115.78,
114.80, 82.93, 65.74, 28.06. MS (EI): m/z Calcd. for C18H20N2O4
[M]+: 328.1423. Found: 328.1420.
d
147.50, 124.43, 114.94, 70.69, 70.52, 79.52, 67.40, 48.64, 47.38, 182
45.81, 8.63. MS (ESI): m/z Calcd. for C82H110N10O20 [M+H]+: 183
Compound 3: Under anhydrous condition, compound 8 (0.3 g,
0.59 mmol), compound 4 (0.58 g, 1.78 mmol), potassium carbon-
ate (0.325 g, 2.36 mmol) and a trace of tetrabutylammonium
bromide were dissolved in dry acetonitrile. The reaction was
refluxed overnight. After cooling to room temperature, and then
filtered. After evaporation solvent, the product was chromato-
graphed on a column of silica (silica gel, 0.5% methanol/CH2Cl2) to
obtain 0.55 g (93%) compound 10 as an orange powder. 1H NMR
1555.7898. Found: 1555.7994.
3. Results and discussion
3.1. Synthesis of hydraphile 1
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Gokel and co-workers designed a series of tri(macrocyle) 187
hydraphiles with different spacer lengths, in which the hydra- 188
philes exhibited incompetent or poor ion transport when the 189
covalent connectors was less than 8 carbon atoms or more than 190
16 carbon atoms in dioleoylphosphatidylcholine (DOPC) bilayer 191
membranes [12h]. The optimal spacer length for hydraphiles in 192
such membranes is 12–16 methylenes. Based on their research, we 193
hoped that replacement of the methylene linker in the tri(macro- 194
(500 MHz, CDCl3):
J = 26.3, 8.5 Hz), 4.85 (d, 4H, J = 6.1 Hz), 4.59 (s, 4H), 3.772–3.575
(m, 24H), 1.56 (s, 18H). 13C NMR (100 MHz, CDCl3):
167.64,
d 7.87 (dd, 8H, J = 8.2, 3.7 Hz), 7.02 (dd, 8H,
d
159.83, 147.50, 124.43, 124.39, 114.94, 114.82, 82.69, 70.68, 69.06,
65.78, 59.17, 28.05, 24.26, 19.81, 13.75. MS (ESI): m/z Calcd. for
C52H66N6O14 [M+H]+: 999.4637. Found: 999.4714.
Compound 2: Under Ar gas protection, compound 3 (0.2 g,
0.2 mmol) was dissolved in 8 mL dry CH2Cl2. Then 2 mL
trifluoroacetic acid was added, and stirred for 3 h at room
temperature. After evaporation solvent, the compound 2 (0.17 g)
cyle) hydraphile with an azobenzene motif would give
a
195
structurally-regulated photosensitive hydraphile channel. As 196
shown in Scheme 1, a bromoacetic acid substituted dihydrox- 197
yazobenzene was selected as the connector due to its similar 198
length to that of 14 carbon chain. It is expected that hydraphile 1 in 199
its trans form would form efficient channel for ion transport, while 200
was obtained. Yield: 96%. 1H NMR (400 MHz, CDCl3):
7.06 (d, 8H, J = 7.3 Hz), 4.99 (s, 4H), 4.75 (s, 4H), 3.69–3.55 (m, 24H).
13C NMR (100 MHz, CDCl3):
169.92, 160.31, 145.54, 124.06,
d 7.82 (s, 8H),
d
Please cite this article in press as: R.-Y. Yang, et al., A light-regulated synthetic ion channel constructed by an azobenzene modified
hydraphile, Chin. Chem. Lett. (2015), http://dx.doi.org/10.1016/j.cclet.2015.05.010

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R.-Y. Yang et al. / Chinese Chemical Letters xxx (2015) xxx–xxx
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the photoisomerization of the hydraphile (in its cis form) would
decrease the transmembrane activity due to the reduced molecular
length equal to that of 10–11 carbon chain. The reversible
photoisomerization, therefore, induced light-regulated ion trans-
port across lipid bilayers. Hydraphile 1 was synthesized in several
simple procedures with high yields (as shown in Scheme 2). All the
compounds were well characterized and confirmed by 1H NMR, 13
NMR and mass spectra.
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3.2. Channel transport assay by HPTS vesicle analysis
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A 8-hydroxy-1,3,6-pyrenetrisulfonate (HPTS) labeled LUV
(150 nm diameter, 10% cholesterol in egg yolk phosphatidylcho-
line (EYPC)) fluorescence assay was carried out to explore the
transport activity of hydraphile 1. In this assay, the ion transport
through the membranes was accessed by the change in ratio
fluorescence intensity at 510 nm (I450/I405) of the pH-sensitive
HPTS dye entrapped inside the vesicles. A pH gradient across the
vesicle membrane was introduced by addition of base (potassium
hydroxide). If the channel mediates ion transport, potassium ions
should flow inside the vesicles, along with OHꢀ (symport) or H+
(antiport) to maintain charge balance. The resultant increase in pH
would cause a deprotonation of the HPTS dye in the vesicles. For
each set of experiments, although the vesicle concentration is not
known precisely, it is constant throughout the experiments. The
concentration-dependent assays were not stopped until the value
of transmembrane activity (Y, ratiometric fluorescence intensity of
HPTS at 350 s) achieved the maximum. Firstly, the solvent effect on
the transmembrane activity was detected, in which DMSO was
used as the blank and showed ignorable activity (ꢁ0.07),
suggesting the successful preparation of vesicles for further assay.
As shown in Fig. 1, addition of hydraphile 1 caused a rapid increase
and followed by a slow and almost linear increase in the
Fig. 1. HPTS assays of hydraphile
concentrations from 0 to 40 mol/L, the presented curves were assigned to DMSO
(blank), 0.037, 0.075, 0.15, 0.6, 2.5, 5.0, 10.0, 20.0, and 40.0 mol/L, respectively.
1
for K+ transport with increasing final
m
m
normalized ratiometric fluorescence of HPTS, suggesting the
successful transport of K+ cations and the rapid partition of the
channel in lipid bilayers [13]. A variation in the concentration of
hydraphile 1 induced a corresponding change in the transmem-
brane activity, displaying efficient transmembrane activity with
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Ymax ꢁ 0.7 at 40
mmol/L of hydraphile 1 (final concentration). The
addition with higher concentration failed to achieve higher
transmembrane activity due to the precipitation from the solution.
When the transmembrane activity observed at 350 s were plotted
as function of concentration, it exhibited a linear relationship at
initial and a flattened tendency for the higher concentrations,
which was indicative of the single-molecular channel transport. All
these suggested that hydraphile 1 formed efficient transmembrane
channel in lipid bilayers and provided the possibility for the further
regulation of ion transport upon light irradiation.
Fig. 2. Evolution of the UV-vis absorption spectra of hydraphile 1 solution (0.2 mmol/L in DMSO) as a function of irradiation time. (a) Upon the irradiation of 365 nm UV light
(5 mW/cm2), t (s) = 0, 5, 10, 15, 20, 25, 30, 35; (b) Upon the irradiation of 450 nm light (5 mW/cm2), t (s) = 0, 3, 6, 9, 12, 18, 22, 30, 40, 60; (c) Switching cycles under alternating
irradiation with 365 nm UV light and 450 nm visible light (2 min each). And (d) the absorbance at 360 nm from (c) procedure.
Please cite this article in press as: R.-Y. Yang, et al., A light-regulated synthetic ion channel constructed by an azobenzene modified
hydraphile, Chin. Chem. Lett. (2015), http://dx.doi.org/10.1016/j.cclet.2015.05.010

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R.-Y. Yang et al. / Chinese Chemical Letters xxx (2015) xxx–xxx
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Fig. 3. The NMR spectra for hydraphile 1 solution (8 mmol/L in CDCl3) before irradiation and after 60 s exposure to UV light (365 nm, 10 mW/cm2).
3.3. Photo-regulated ion transport
(trans:cis) was observed by NMR. All these indicated the reversible
photo-regulation on the molecular structure.
Before investigating the light-regulated action for the ion
transport by the isomerization of azobenzene group, the fact that
trans-azobenzene was indeed reversibly isomerized to the cis-
isomer was confirmed firstly by UV–vis spectroscopic studies in
the solution state. As shown in Fig. 2a, upon 365 nm UV irradiation
and with an increase in irradiation time, the absorption band at
360 nm decreased, which was accompanied by the appearance of a
band at 450 nm, indicating the photo-isomerization of hydraphile
1 from its trans form to cis form. The generation of isosbestic points
at 320 nm and 430 nm indicated the clean photoisomerization
The photo-regulated ion transport of the molecules in BLMs was
also investigated by HPTS assays. A 365 nm and a 450 nm LED
lamps with an intensity of 30 mW/cm2 were used to irradiate the
DMSO stock solutions containing hydraphile 1. In this experiment,
to ensure the obtained ion transport was induced from trans- or cis-
isomer, the DMSO solution of hydraphile 1 was irradiated with
corresponding light for minutes prior to addition to the vesicle
suspension in the assay. To optimize the photo-regulation of
transmembrane activity, 5
mmol/L hydraphile 1 (final concentra-
tion) was applied. As illustrated in Fig. 4a, the transmembrane
activity of hydraphile 1 was around 0.45 before any irradiation. It
was noticed that 10% cis-isomer was already present in the fresh
prepared compound, irradiation with 450 nm light was firstly
performed in order to detect the transmembrane activity of trans-
isomer. As expected, the transmembrane activity was increased to
0.5 after 10 min pre-irradiation with 450 nm light. Then the DMSO
stock solution was irradiated with 365 nm light for 10 min, and
the transmembrane activity decreased to 0.35. It indicated that
reaction for hydraphile
1 upon irradiation. Conversely, the
irradiation with 450 nm visible light induced the recovery of the
trans-isomer and the trans–cis cycle was reversible under
alternating irradiation with 365 nm UV light and 450 nm visible
light (as shown in Fig. 2b and d). The conversion ratio of trans- to
cis-isomers could be determined by the corresponding proton NMR
studies as illustrated in Fig. 3. Selective irradiation at 365 nm
increased the amount of cis-isomer and a final ration of 10:90
Fig. 4. (a) Observed change in transmembrane activity of HPTS assay of hydraphile 1 (5.0
mmol/L, final concentration) upon the irradiation of light-365 nm UV light or 450 nm
visible light, the middle one is the original activity without any irradiations; (b) cycles of transmembrane activity under alternating irradiation with 365 nm UV light and
450 nm visible light (5 min each). Irradiation condition: 365 nm UV light and 450 nm visible light, intensity: 10 mW/cm2.
Please cite this article in press as: R.-Y. Yang, et al., A light-regulated synthetic ion channel constructed by an azobenzene modified
hydraphile, Chin. Chem. Lett. (2015), http://dx.doi.org/10.1016/j.cclet.2015.05.010

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[5] (a) M.R. Banghart, M. Volgraf, D. Trauner, Engineering light-gated ion channels,
Biochemistry 45 (2006) 15129–15141;
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trans-isomer of hydraphile 1 provided higher transmembrane
activity than that of cis-isomer. Meanwhile, the transmembrane
activity showed reversible regulation upon alternative irradiation
of 365 nm UV light and 450 nm visible light (as shown in Fig. 4b).
All these verified the intention for the initial design of light-
regulated ion channel by reversible regulation of the transmem-
brane length of molecule. Although the obtained regulation
between trans and cis-isomers was relative small (transmembrane
activity decreased 0.15 from trans to cis), it was assured that
excellent light-regulated ion channel could be constructed by
further molecular optimization in future.
(b) T. Fehrentz, M. Scho¨nberger, D. Trauner, Optochemische genetik, Angew.
Chem. Int. Ed. 50 (2011) 12156–12182;
(c) M.R. Banghart, A. Mourot, D.L. Fortin, et al., Photochromic blockers of voltage-
gated potassium channels, Angew. Chem. Int. Ed. 48 (2009) 9097–9101;
(d) M. Volgraf, P. Gorostiza, R. Numano, et al., Allosteric control of an ionotropic
glutamate receptor with an optical switch, Nat. Chem. Biol. 2 (2006) 47–52.
[6] (a) A. Koc¸er, M. Walko, W. Meijberg, B.L. Feringa, A light-actuated nanovalve
derived from a channel protein, Science 309 (2005) 755–758;
´
(b) W. SzymaNski, D. Yilmaz, A. Koc¸er, B.L. Feringa, Bright ion channels and lipid
bilayers, Acc. Chem. Res. 46 (2013) 2910–2923;
(c) C. Bao, H. Jia, T. Liu, Y. Wang, W. Peng, L. Zhu, Synthesis of artificial ion
channels in bilayer membrane, Prog. Chem. 24 (2012) 1337–1345.
[7] (a) P.V. Jog, M.S. Gin, A light-gated synthetic ion channel, Org. Lett. 10 (2008)
3693–3696;
(b) P. Osman, S. Martin, D. Milojevic, C. Tansey, F. Separovic, Optical modulation
of the insertion of gramicidin into bilayer lipid membranes, Langmuir 14 (1998)
4238–4242.
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4. Conclusion
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268
269
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271
272
273
274
275
276
In summary, we have demonstrated a new type of light-
regulated synthetic ion channel based on azobenzene substituted
tri (macrocycle) hydraphile 1. HPTS vesicle assay confirmed the
efficient transport of hydraphile 1 for ion across the lipid bilayers.
Photoisomerization of the azobenzene using 365 nm UV light and
450 nm visible light resulted in the attenuation of channel
transmembrane activity. Work is ongoing to create and develop-
ment of these robust synthetic gated ion channels, which would
exhibit great potential in various nanotechnology and biomechan-
ical applications.
[8] (a) V. Borisenko, D.C. Burns, Z. Zhang, G.A. Woolley, Optical switching of ion-
dipole interactions in a gramicidin channel analogue, J. Am. Chem. Soc. 122 (2000)
6364–6370;
(b) L. Lien, D.C.J. Jaikaran, Z. Zhang, G.A. Woolley, Photomodulated blocking of
gramicidin ion channels, J. Am. Chem. Soc. 118 (1996) 12222–12223.
[9] L. Husaru, R. Schulze, G. Steiner, et al., Potential analytical applications of gated
artificial ion channels, Anal. Bioanal. Chem. 382 (2005) 1882–1888.
[10] (a) A.A. Beharry, G.A. Wolley, Azobenzene photoswitches for biomolecules,
Chem. Soc. Rev. 40 (2011) 4422–4437;
(b) D.G. Flint, J.R. Kumita, O.S. Smart, G.A. Woolley, Using an azobenzene cross-
linker to either increase or decrease peptide helix content upon trans-to-cis
photoisomerization, Chem. Biol. 9 (2002) 391–397;
(c) M.L. Rahman, G. Hegde, S.M. Sarkar, M.M. Yusoff, Synthesis and photoswitch-
ing properties of azobenzene liquid crystals with a pentafluorobenzene terminal,
Chin. Chem. Lett. 25 (2014) 1611–1614.
277
Acknowledgments
[11] (a) T. Liu, C. Bao, H. Wang, et al., Light-controlled ion channels formed by
amphiphilic small molecules regulate ion conduction via cis–trans photoisome-
rization, Chem. Commun. 49 (2013) 10311–10313;
278 Q3
279
280
This work was supported by NNSFC (Nos. 51273064,
21472044), Innovation Program of Shanghai Municipal Education
Commission and Fundamental Research Funds for the Central
University.
(b) T. Liu, C. Bao, H. Wang, et al., Self-assembly of crown ether-based amphiphiles
for constructing synthetic ion channels: the relationship between structure and
transport activity, New J. Chem. 38 (2014) 3507–3513;
(c) C.L. Murray, G.W. Gokel, Spacer chain length dependence in hydraphile
channels: implications for channel position within phospholipid bilayers, J.
Supramol. Chem. 1 (2001) 23–30.
281
282
References
[12] (a) G.W. Gokel, S. Negin, Synthetic ion channels: from pores to biological appli-
cations, Acc. Chem. Res. 46 (2013) 2824–2833;
283
284
285
286
287
288
289
290
291
292
293
294
295
296
297
298
299
300
301
302
303
[1] (a) C. Brieke, F. Rohrbach, A. Gottschalk, G. Mayer, A. Heckel, Light-controlled
tools, Angew. Chem. Int. Ed. 51 (2012) 8446–8476;
(b) A. Nakano, Q. Xie, J.V. Mallen, L. Echegoyen, G.W. Gokel, Synthesis of a
membrane-insertable, sodium cation conducting channel: kinetic analysis by
dynamic 23Na NMR, J. Am. Chem. Soc. 112 (1990) 1287–1289;
(c) G.W. Gokel, Hydraphiles: design, synthesis and analysis of a family of syn-
thetic, cation-conducting channels, Chem. Commun. 1 (2000) 1–9;
(d) C.L. Murray, H. Shabany, G.W. Gokel, The central ‘relay’ unit in hydraphile
channels as a model for the water- and-ion ‘capsule’ of channel proteins, Chem.
Commun. 23 (2000) 2371–2372;
(e) M.E. Weber, P.H. Schlesinger, G.W. Gokel, Dynamic assessment of bilayer
thickness by varying phospholipid and hydraphile synthetic channel chain
lengths, J. Am. Chem. Soc. 127 (2005) 636–642;
(f) O. Murillo, I. Suzuki, E. Abel, et al., Synthetic transmembrane channels:
functional characterization using solubility calculations, transport studies, and
substituent effects, J. Am. Chem. Soc. 119 (1997) 5540–5549;
(g) W.M. Leevy, G.M. Donato, R. Ferdani, et al., Synthetic hydraphile channels of
appropriate length kill Escherichia coli, J. Am. Chem. Soc. 124 (2002) 9022–
9023;
(b) A.A. Beharry, G.A. Woolley, Azobenzene photoswitches for biomolecules,
Chem. Soc. Rev. 40 (2011) 4422–4437;
(c) D. Habault, H. Zhang, Y. Zhao, Light-triggered self-healing and shape-memory
polymers, Chem. Soc. Rev. 42 (2013) 7244–7256.
[2] (a) W. Szyman´ ski, J.M. Beierle, H.A.V. Kistemaker, W.A. Velema, B.L. Feringa,
Reversible photocontrol of biological systems by the incorporation of molecular
photoswitches, Chem. Rev. 113 (2013) 6114–6178;
(b) R. Givens, M.B. Kotala, J.I. Lee, Dynamic Studies in Biology, Wiley-VCH Verlag
GmbH & Co. KGaA, 2005, pp. 95–129;
(c) P. Gorostiza, E. Isacoff, Optical switches and triggers for the manipulation of
ion channels and pores, Mol. BioSyst. 3 (2007) 686–704.
[3] G. Nagel, D. Ollig, M. Fuhrmann, et al., Conversion of channelrhodopsin into a
light-gated chloride channel, Science 296 (2002) 2395–2398.
[4] (a) R.J. Thompson, M.F. Jackson, M.E. Olah, et al., Activation of pannexin-1
hemichannels augments aberrant bursting in the hippocampus, Science 322
(2008) 1555–1559;
(h) B.A. Smith, M.M. Daschbach, S.T. Gammon, et al., In vivo cell death mediated
by synthetic ion channels, Chem. Commun. 47 (2011) 7977–7979.
[13] C.P. Wilson, C. Boglio, L. Ma, S.L. Cockroft, S.J. Webb, Palladium(II)-mediated
assembly of biotinylated ion channels, Chem. Eur. J. 17 (2011) 3465–3473.
(b) S. Sundelacruz, M. Levin, D.L. Kaplan, Role of membrane potential in the
regulation of cell proliferation and differentiation, Stem Cell Rev. Rep. 5 (2009)
231–246.
Please cite this article in press as: R.-Y. Yang, et al., A light-regulated synthetic ion channel constructed by an azobenzene modified
hydraphile, Chin. Chem. Lett. (2015), http://dx.doi.org/10.1016/j.cclet.2015.05.010