Preorganized aryltriazole foldamers as effective transmembrane transporters for chloride anion

Article abstract of DOI:10.1021/ol501772v

Preorganized aryltriazole foldamers 1 and 2 were designed and synthesized. NMR studies and X-ray analysis demonstrate that 1 adopts a crescent conformation driven by a series of continuous hydrogen bonds at the periphery of the foldamer, whereas 2 displays a coil conformation. NMR titrations reveal that the affinities of fully preorganized foldamer 1 for halogen anions are much stronger that those of partially preorganized foldamer 2. Furthermore, it is found that such full preorganization makes 1 an effective transmembrane transporter for the chloride anion across a lipid bilayer.

Full text of DOI:10.1021/ol501772v

Letter
pubs.acs.org/OrgLett
Preorganized Aryltriazole Foldamers as Effective Transmembrane
Transporters for Chloride Anion
Jie Shang,†,§ Wen Si,‡ Wei Zhao,†,§ Yanke Che,§ Jun-Li Hou,*,‡ and Hua Jiang*,†
†College of Chemistry, Beijing Normal University, Beijing 100875, China
‡Department of Chemistry, Fudan University, 220 Handan Road, Shanghai 200433, China
§CAS Key Laboratory of Photochemistry, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190, China
S
* Supporting Information
ABSTRACT: Preorganized aryltriazole foldamers 1 and 2 were designed and synthesized. NMR studies and X-ray analysis
demonstrate that 1 adopts a crescent conformation driven by a series of continuous hydrogen bonds at the periphery of the
foldamer, whereas 2 displays a coil conformation. NMR titrations reveal that the affinities of fully preorganized foldamer 1 for
halogen anions are much stronger that those of partially preorganized foldamer 2. Furthermore, it is found that such full
preorganization makes 1 an effective transmembrane transporter for the chloride anion across a lipid bilayer.
transporters have been developed so as to shed light into the
eveloping synthetic receptors for selectively binding
Danions has attracted increasing attention in the past few mechanisms of these genetic diseases and also to provide
potential therapies for treating them.13 However, examples of
foldamer-based anion transporters are extremely rare.14
Previously, we and others reported a series of chloride
mediated aryltriazole foldamers.10,11 Later, we successfully
replaced some triazole moieties with more acidic amides so
that the affinities of aryltriazole foldamers for halogen anions
were significantly enhanced.11c On the other hand, Jeong9 and
Flood10 utilized an alternative approach to achieve the same
purpose by partially rigidifying backbones of foldamers.
Encouraged by these results, herein, we report our design
and syntheses of the preorganized aryltriazole foldamers
(Figure 1) and their functions as transmembrane transporters
for chloride. To our delight, we found that the full
preorganization makes aryltriazole foldamer 1 a potent anion
chelator and an effective transmembrane transporter for
chloride anions across a lipid bilayer.
decades because of their biological importance.1 Foldamers,2
artificial analogues of biopolymers having strong a tendency to
adopt well-defined conformation, hold great promise as hosts
for biologically interesting guests because of their dynamic
internal cavities or channels. Prominent examples for foldamers
serving as hosts include m-phenylene ethynylene oligomers,3
oligo(m-ethynylpyridine)s,4 hydrogen-bonded oligohydrazide
foldamers,5 capsule-like oligoamide foldamers,6 oligocholates
and metallohelicenes,7,8 oligoindole-based foldamers, and
aryltriazole foldamers.9−11 However, it is true to say that the
systems of foldamer-based anion recognitions are not fully
developed yet, despite the fact that a number of synthetic anion
receptors1 and foldamers adopting well-defined helical
conformations2 have been reported so far. The challenge in
the field of anion-mediated foldamers is how to efficiently
synthesize elegant foldamers with suitable anion-binding sites in
the internal cavity.
We have long-term interest in foldamer-based molecular
recognitions,6 particularly in anion recognitions.11 This is
because anions play vital roles in biological processes.1 It is
believed that the metabolic dysregulation of anions is
implicated to a number of genetic diseases including most
notably cystic fibrosis and the renal disease Bartter’s
syndrome.12 Therefore, a number of transmembrane anion
The N2/N3 atoms in triazole provide ideal hydrogen bond
acceptors. The formation of hydrogen bonds with the triazole’s
N3 atom were recently demonstrated by Flood.10 On the other
hand, the NH in amide is widely used as a hydrogen bond
donor in the construction of foldamers. Accordingly, we
Received: June 25, 2014
© XXXX American Chemical Society
A
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Letter
bonded structure. Such a series of continuous hydrogen bonds
at the periphery of foldamer block free rotation of triazoles and
aromatic rings. In such a situation, only NOE correlations
between triazole proton Hc and adjacent protons Hb and Hd
are observable (SI). These NMR data further confirm that 1
adopts a crescent conformation as observed in the solid phase.
However, in the case of 2, although similar hydrogen bonds
between amides and triazoles are observed (Figure 3B), such
hydrogen bonds are unable to fully rigidify the backbone of 2.
The free rotations between the triazole moieties and adjacent
aromatic rings were confirmed by 2D NOESY experiment (SI),
implying an unfolded conformation for 2.
The NMR titrations of 1 with the chloride anion as shown in
Figure 3A are representative of the features observed for Cl−,
Br− and I−. The titrations of chloride anions cause all of the
inner protons expected to bind chloride to significantly shift
downfield by great amounts (SI). For example, the proton Hc
shifts downfield by the largest amount up to 2.21 ppm (Figure
3C and Table S1, SI). In addition, the chemical shifts of the
protons of amides display slight variations upon the titrations of
halide anions, indicating that the protons in the cavity primarily
serve in binding the halide anions. In the case of bromide and
iodide anion titrations, similar but weaker downfield shifts than
those in the case of chloride were observed for all of the inner
protons. The largest downfield shifts of the proton Hc are 2.06
and 1.70 ppm for Br− and I−, respectively (Figure 3C and SI).
The binding stoichiometries for all anions were determined to
be 1:1 by Job’s plot (SI). The binding constants of 1 for Cl−,
Br− and I− anions were calculated on the basis of the NMR
titrations by the WinEQNMR program15 to be 757, 367, and
134 M−1, respectively (Table S1, SI). The binding affinities (K)
and largest changes in chemical shifts (Δλ) of Hc appear in the
order of Cl− > Br− > I− as a result of the electrostatic nature of
hydrogen-bonding interaction.
Figure 1. Structures of preorganized foldamers 1 and 2.
designed pentad 1, in which the amide moieties were
introduced into aromatic rings adjacent to triazoles so as to
form four intramolecular hydrogen bonds with the triazoles’
N2/N3 atoms, respectively. We expected that these intra-
molecular hydrogen bonds would eventually result in a full
backbone-rigidified foldamer with enhanced affinity for halogen
anion. In addition, a partial preorganized foldamer 2 has also
been designed for comparison. The synthetic routes and full
characterization data for 1 and 2 are provided in the Supporting
Information (SI).
The preorganization in the aryltriazole foldamer 1 was first
confirmed by the X-ray analysis. Single crystals of 1 were
obtained by evaporating its solution in dichloromethane/ethyl
acetate. As expected, each triazole is held by two intramolecular
hydrogen bonds between the amides NH and the triazole’s N2/
N3 atoms, respectively, as shown in Figure 2. However, the
In the case of 2, the representative NMR titration spectra are
shown in Figure 3B. The tendencies in the variations of the
chemical shifts of 2 are similar to those of 1 upon the addition
of halide anions. The protons of triazoles significantly move
downfield. For example, Hh shifts downfield by the amount of
1.80, 1.69, and 1.35 ppm for Cl−, Br−, and I−, respectively (SI).
The affinities of 2 for Cl−, Br− and I− were extracted from
NMR data to be 91, 109, and 86 M−1, respectively (Table S1,
SI). These values are much smaller than those in the case of 1
and appear in the order of Br− > Cl−> I− in the contrary to the
foldamer 1. The differences in the order presumably are caused
by the flexible backbone of 2, which is just right for Br−. The
largest difference in the affinity is found for the chloride anion
up to about 8 times. Such differences in the affinities
presumably originate from the extent of preorganization in
the backbones of ary-triazole foldamers.
The halogen anion-binding properties of 1 and 2 prompted
us to investigate the capacity of these molecules of transporting
halogen anions across lipid bilayer. Thus, the Cl− transport
behaviors of the new aryltriazole foldamers 1 and 2 were first
studied by imposing a Cl− gradient across the lipid membrane
of the large unilamellar vesicles (LUVs) made from the egg
yolk L-α-phosphatidylcholine (EYPC). A suspension of vesicles
entrapping the Cl−-sensitive dye lucigenin was added to a buffer
containing KCl to produce a KCl gradient.16 The transport of
Cl− into the vesicles was assessed by monitoring the
fluorescence intensity of lucigenin. Adding 1 and 2 [molar
ratio relative to lipid (x) = 0.75%] to the above vesicle
suspension caused a small change (25%) in fluorescence
Figure 2. Top and side views of the crystal structure of 1. The
distances of hydrogen bonds were labeled and highlighted in green.
The protons in tert-butyl groups were deleted for clarity.
distances of hydrogen bonds involved in one triazole (about
2.05 Å) are a little longer than those in the other, leading to the
tilt-up of one triazole. In addition, four weak hydrogen bonds
between the amide’s carbonyl groups and adjacent CHs in the
aromatic rings are also observed. Taken together, the backbone
of aryltriazole foldamer 1 is greatly rigidified by these hydrogen
bonds, which consequently results in the terminal aromatic
rings tilting up and down according to the central aromatic ring
(side view in Figure 2). Efforts on obtaining crystals of 2 and
complexes 1TBACl and 2TBACl failed.
The labeled protons and solution structures of 1 and 2 were
assigned and characterized by HSQC, HMBC and 2D NOESY
experiments in CD2Cl2 and CDCl3, respectively (see the SI). In
the case of 1, as shown in Figure 3A, the amide protons are
deshielded at 10−11.5 ppm, as is expected for a hydrogen-
B
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1
Figure 3. Changes in the 1H chemical shift of 1 and 2. (A) Partial H NMR (298 K, 400 MHz) spectral changes of 1 in CD2Cl2 upon addition of
TBACl. (B) Partial 1H NMR spectral changes of 2 in CDCl3 upon addition of TBACl. (C) Changes in 1H chemical shift for the triazole (Hc) of 1 in
CD2Cl2 with increasing concentrations of different anions. (D) Changes in 1H chemical shift for the triazole (Hh) of 2 in CDCl3 with increasing the
various concentrations of different anions; [1] = [2] = 2 mM.
intensity (Figure 4a). However, adding their mixtures with
the present case. Our results demonstrate that the affinities of
the foldamers are just right for effective transport. The
transport experiments of 1 and 2 for Br− and I− were also
conducted by using the same methods. However, the transport
was not measurable, since Br− and I− can pass through the lipid
bilayer.
valinomycin (0.002 mol % relative to lipid), an efficient K+
It has been established that ion transmembrane transport can
be achieved via carrier13b or channel mechanism.18 To further
investigate the transport mechanism, Cl− transport experiments
were also carried out by using LUVs composed of EYPC and
cholesterol (2:1 in molar ratio). It has been reported that
cholesterol decreases the fluidity of a lipid bilayer19 and that
this would have a negative effect on the transport ability of a
carrier,20 which is largely dependent on movement within the
bilayer. The Cl-transport activities of 1 and 2 in 2:1
EYPC:cholesterol vesicles show a moderate decrease in
comparison with these in EYPC vesicles under the same
conditions (see the SI), implying the mobile carrier mechanism.
The patch clamp experiments on planar lipid bilayer were also
performed. By premixing 1 or 2 with the lipids and painting to
form a planar lipid bilayer, no conductance signal was observed,
which further confirms the carrier mechanism.
To quantitatively measure the activity of carrier 1, transport
experiments were also carried out under different carrier
concentrations. It was found that the transport ability, defined
as the relative fluorescence intensity of lucigenin after 12 min of
adding 1, was strongly dependent on x. As x increased from 0
to 0.2%, the intensity increased significantly. Further increasing
x caused only a slight increase in intensity (Figure 5). The
effective concentration needed for achieving 50% (EC50)
transport ability and the Hill coefficient (n) were calculated
to be 0.09% and 1.3,21 respectively. Such a low EC50 indicates
that the new foldamer is an efficient transporter for Cl−. The
Hill coefficient value closes to a unit, demonstrating that 1
transports Cl− in a single-molecular-manner.21
Figure 4. (a) Changes in fluorescence intensity (λex = 372 nm, λem
=
503 nm) of vesicles with time after addition of 1 or 2 (x = 0.75%)
and/or valinomycin (x = 0.002%). The vesicles loaded with lucigenin
(2.0 mM) buffered at pH 7.0 with HEPES (10 mM) and suspended in
KCl (100 mM) solution (buffered to pH 7.0). (b, c) Schematic
representation of blockage and permit transport of Cl− in the absence
and presence of valinomycin (V).
carrier,17 caused the intensity to increase by 97% and 39%,
respectively, within 12 min. In contrast, the same amount of
valinomycin itself resulted in only an 11% increase in
fluorescence intensity. These results support the conclusion
that 1 and 2 can transport Cl−. The transport ability can be
obviously enhanced by valinomycin. The enhancement of 1 is
more remarkable than that of 2, demonstrating that 1 exhibits
higher transport ability. This is not unexpected because 1
possesses higher affinity to Cl− than 2 does. Considering that 1
and 2 have a similar backbone, it is reasonable to believe that
this is a result of their differences in binding affinities to Cl−,
although the better binding does not always lead to effective
transport, which is the so-called “Goldilocks principle”. Such a
phenomenon was observed in the Matile’s system13g but not in
In summary, the conformations of aryltriazole foldamers can
be preorganized by the incorporation of hydrogen bonds at the
periphery of foldamers, but the extent of preorganization is
greatly dependent on the number of hydrogen bonds, which
poses a great effect on the affinity of foldamer for anions. The
C
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ASSOCIATED CONTENT
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* Supporting Information
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Synthetic procedures, H NMR, 13C NMR, and 2D NMR
spectra of 1 and 2 and X-ray crystallographic data. This material
is available free of charge via the Internet at http://pubs.acs.org.
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AUTHOR INFORMATION
Corresponding Authors
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*E-mail: houjl@fudan.edu.cn.
*E-mail: jiangh@bnu.edu.cn.
Notes
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P. A.; Iglesias-Sanchez, J. C.; Prados, P.; Quesada, R. J. Am. Chem. Soc.
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The authors declare no competing financial interest.
ACKNOWLEDGMENTS
We thank the National Natural Science Foundation of China
(21125205 and 21332008).
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