Rotaxanes and biofunctionalized pseudorotaxanes via thiol-maleimide click chemistry

Article abstract of DOI:10.1021/ol300614z

Base-catalyzed thiol-maleimide click chemistry has been applied to the synthesis of neutral donor-acceptor [2]rotaxanes in good yield. This method is extended further to the synthesis of a glutathione-functionalized [2]pseudorotaxane, a precursor to integ

Full text of DOI:10.1021/ol300614z

ORGANIC
LETTERS
2012
Vol. 14, No. 8
2082–2085
Rotaxanes and Biofunctionalized
Pseudorotaxanes via Thiol-Maleimide
Click Chemistry
Umesh Choudhary and Brian H. Northrop*
Department of Chemistry, Wesleyan University, Middletown, Connecticut 06459,
United States
bnorthrop@wesleyan.edu
Received March 10, 2012
ABSTRACT
Base-catalyzed thiol-maleimide click chemistry has been applied to the synthesis of neutral donorꢀacceptor [2]rotaxanes in good yield. This
method is extended further to the synthesis of a glutathione-functionalized [2]pseudorotaxane, a precursor to integrated conjugates of interlocked
molecules with proteins and enzymes.
Molecules possessing mechanical bonds have attracted
significant research interest for more than five decades.1
Initial syntheses of mechanically interlocked catenanes
and rotaxanesbyWasserman2 and Harrison,3 respectively,
were achieved in very low yield (<5%) owing to the sta-
tistical formation of interlocking species. Beginning in the
1980s, however, synthetic methods for the preparation of
mechanically interlocked molecules have advanced apace,
largely through the use of template-directed self-assembly
protocols4 used in conjunction with kinetic and thermo-
dynamic covalent bond forming reactions5 that “lock”
individual components together. Increased synthetic effi-
ciency has allowed increasingly complex mechanically
interlocked structures to be synthesized, many of which
have been designed to be stimuli-responsive6 and form the
basis of various molecular machines.7 Toward the aim of
increasing both synthetic ease and structural complexity,
(6) For representative examples of interlocked molecules that can be
stimulated electrochemically, see: (a) Green, E.; Choi, J. W.; Boukai, A.;
Bunimovich, Y.; Johnston-Halprin, E.; DeIonno, E.; Luo, Y.; Sheriff,
B. A.; Xu, K.; Shin, Y. S.; Tseng, H.-R.; Stoddart, J. F.; Heath, J. R.
Nature 2007, 445, 414–417. Stimulated with acidꢀbase inputs: (b)
Badjic, J. D.; Balzani, V.; Credi, A.; Stoddart, J. F. Science 2004, 303,
1845–1849. Stimulated photolytically: (c) Leigh, D. A.; Wong, K. Y.;
Dehez, F.; Zerbetto, F. Nature 2003, 424, 174–179. (d) Serreli, V.; Lee,
C.-F.; Kay, E. R.; Leigh, D. A. Nature 2007, 445, 523–527. Stimulated
using alkali or transition metal coordination: (e) Collin, J.-P.; Dietrich-
(1) (a) Catenanes, Rotaxanes, and Knots; Sauvage, J.-P., Dietrich-
Buchecker, C., Eds.; Wiley-VCH: Weinheim, 1999. (b) Stoddart, J. F. Chem.
Soc. Rev. 2009, 38, 1802–1820. (c) Olsen, J.-C.; Griffiths, K. E.;
Stoddart, J. F. A Short History of the Mechanical Bond. In From
Non-Covalent Assemblies to Molecular Machines; Sauvage, J.-P., Gaspard, P.,
Eds.; Wiley-VCH: Weinheim, 2011; pp 67ꢀ139.
(2) Wasserman, E. J. Am. Chem. Soc. 1960, 82, 4433–4434.
(3) Harrison, I. T.; Harrison, S. J. Am. Chem. Soc. 1967, 89, 5723–
5724.
~
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Wiley-VCH: Weinheim, 2000. (b) Busch, D. H. Top. Curr. Chem. 2005,
249, 1–65.
Buchecker, C.; Gavina, P.; Jimenez-Molero, M. C.; Sauvage, J.-P. Acc.
Chem. Res. 2001, 34, 477–487. (f) Vignon, S. A.; Jarrosson, T.; Iijima, T.;
Tseng, H.-R.; Sanders, J. K. M.; Stoddart, J. F. J. Am. Chem. Soc. 2004,
126, 9884–9885.
ꢁ
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K.; Aprahamian, I.; Heath, J. R.; Stoddart, J. F. Acc. Chem. Res. 2008,
41, 1750–1761.
(7) Kay, E. R.; Leigh, D. A.; Zerbetto, F. Angew Chem. Int. Ed. 2006,
46, 72–191 and references therein.
r
10.1021/ol300614z
Published on Web 04/06/2012
2012 American Chemical Society

Scheme 1. Noncovalent Self-Assembly of [2]Pseudorotaxane 4 Followed by Thiol-Maleimide Click Chemistry Mediated Interlocking
To Give [2]Rotaxane 5 in the Presence of Catalytic Et3N
one of the most notable advances in the construction of
mechanically interlocked molecules has been the introduc-
tion of highly efficient “click” reactions to their synthesis.8
In 2006, the groups of Leigh,9 Sauvage,10 and Stoddart11
each independently reported the efficient synthesis of inter-
locked rotaxanes utilizing copper(I) catalyzed alkyneꢀ
azide click (CuAAC) chemistry,12 obtaining [2] and
[3]rotaxanes in up to 94% isolated yield.9
The many attributes of click reactions, particularly the
ability to tolerate low temperatures and high concentra-
tions, make them particularly well suited to the threading
followed by stoppering approach to mechanically inter-
locked molecules, as the thermodynamically controlled
threading step is most favorable at low temperatures and
high concentrations. While the vast majority of click
chemical approaches to mechanically interlocked synthesis
have used the CuAAC reaction, other click methods
such as those using thiol-yne,13 thiol-ene,14 and nitrile-N-
oxide15 procedures have been successfully applied as well.
Here we report the application of thiol-maleimide click
chemistry16 to the synthesis of mechanically interlocked
rotaxanes and noncovalently associated pseudorotaxanes.
The highly selective reactivity of thiols for maleimides has
been utilized for many years in the context of bioconjugate
chemistry.17 In the presence of catalytic base the thiol-
maleimide Michael addition product can be obtained
rapidly (minutes) and often in quantitative yields. Indeed
the extensive history of thiol-maleimide bioconjugate
chemistry is what inspired us to explore the use of thiol-
maleimide chemistry in the synthesis of interlocked mole-
cules. Once suitable conditions are developed it is expected
that stimuli-responsive mechanically interlocked mole-
cules can be directly integrated with biological systems
(e.g., proteins, enzymes) through thiol-maleimide click
reactions involving maleimide-functionalized supramole-
cules and free thiol moieties of solvent assessible cysteine
residues. Toward this aim we demonstrate the efficient
synthesis and self-assemblyof a glutathione-functionalized
[2]pseudorotaxane. Such integration of artificial molecular
machines with biological systems may open the door to
external allosteric control of the functionality of biological
molecules.
The design of maleimide-functionalized naphthalene
diimide (NDI) guest 1 and thiol-functionalized sterically
imposing “stopper” 2 is shown in Scheme 1. Noncovalent
self-assembly of the electron poor naphthalene diimide
with electron rich crown ethers such as DNP38C10
has been well studied,18 particularly by Sanders6f,19 and
co-workers. The interaction between NDI derivatives and
(8) For two recent reviews of the use of click chemistry in the
€
synthesis of mechanically interlocked molecules, see: (a) Hanni,
K. D.; Leigh, D. A. Chem. Soc. Rev. 2010, 1240–1251. (b) Fahrenbach,
A. C.; Stoddart, J. F. Chem.;Asian J. 2011, 6, 2660–2669.
DNP38C10 (Ka ≈ 102 M
)
ꢀ1 19h is not typically as strong as
the interactions between dibenzylammonium guests with
€
(9) Aucagne, V.; Hanni, K. D.; Leigh, D. A.; Lusby, P. J.; Walker,
D. B. J. Am. Chem. Soc. 2006, 128, 2186–2187.
(10) Mobian, P.; Collin, J.-P.; Sauvage, J.-P. Tetrahedron Lett. 2006,
47, 4907–4909.
(17) Brinkley, M. Bioconjugate Chem. 1992, 3, 2–13 and references
therein.
ꢁ
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Stoddart, J. F. J. Am. Chem. Soc. 2006, 128, 10388–10390.
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Klivansky, L. M.; Cao, D.; Snauko, M.; Teat, S. J.; Struppe, J. O.; Liu, Y.
J. Am. Chem. Soc. 2009, 131, 2078–2079. (f) Cao, D.; Amelia, M.;
Klivansky, L. M.; Koshkakaryan, G.; Khan, S. I.; Semeraro, M.; Silvi,
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(g) Yang, W.; Li, Y.; Ahang, J.; Chen, N.; Chen, S.; Lui, H.; Li, Y. J. Org.
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1573 and references therein.
Org. Lett., Vol. 14, No. 8, 2012
2083

instantaneous formation of a deep red solution. 1H NMR
spectroscopy of this solution (Figure 1C) indicated the
formation of thermodynamically stable[2]pseudorotaxane
4 through changes in the chemical shifts of diagnostic
proton signals Hb-He. At 300 MHz and 298 K dethreading
of macrocycle 3 onto and off of thread 1 is comparable
1
to the H NMR time scale, leading to significant broad-
ening of resonances for bothcompounds. A better resolved
spectrum of [2]pseudorotaxane 4 is obtained at 263 K
(Figure 1C) where the dethreading rate is decreased.
Further evidence of a self-assembling hostꢀguest complex
is obtained from UV/vis spectroscopy (see Supporting
Information). A prominent charge-transfer (CT) band is
observed at 487 nm for a 0.001 M CHCl3 solution of 1 and
3 (ε = 143 Mꢀ1 cmꢀ1).
Adding 2.2 equiv of thiol-functionalized stopper 2 to a
0.1 M CDCl3 solution of [2]pseudorotaxane 4 at 273 K in
the presence of 0.03 equiv of Et3N results in the formation
of mechanically interlocked [2]rotaxane 5 (Scheme 1B) in
65% isolated yield. The formation of mechanically inter-
locked [2]rotaxane 5 was confirmed (see Supporting Infor-
mation) by accurate mass APCI mass spectrometric ana-
lysis: m/z = 2653.28 [M þ Na]þ and 1338.13 [M þ 2Na]2þ
compared with calculated values of 2653.28 and 1338.14,
respectively. It is interesting to note that the relative
intensity of the [M þ 2Na]2þ peak of [2]rotaxane 5 is twice
that of the [M þ Na]þ peak. Super- and supramolecular
assemblies of NDI derivatives with DNP38C10 are
known6f,19h to interact with alkali metals in solution. Mass
spectrometric analysis of [2]rotaxane 5 suggests these
interactions persist in the gas phase as well. Mass spectra
of the macrocycle and thread components in isolation
show weak relative intensities of [M þ 2Na]2þ peaks (see
Supporting Information).
Figure 1. Partial 1H NMR spectra (298 K unless otherwise
noted) of thread 1 (A), macrocycle 3 (B), [2]pseudorotaxane 4
(C) (263 K), and [2]rotaxane 5 (D). Protons are labeled as in
Scheme 1. Uncomplexed peaks are designated as “(uc)”.
dibenzo-24-crown-8 hosts (Ka = 102ꢀ104 M
)
ꢀ1 20 or those
between bipyridinium derivatives with DNP38C10 hosts
(Ka = 104ꢀ105 Mꢀ1).21 NDI derivatives are, however,
stable to base and therefore suitable for base catalyzed
thiol-maleimide reactions whereas dialkylammonium
and N-benzylbipyridinium guests are susceptible to
deprotonation6b or nucleophilic attack,22 respectively.
Mixing a 1:1 molar ratio of 1 and DNP38C10 macro-
cycle 3 in CHCl3 at ambient temperature results in the
The 1H NMR spectrum of 5 (Figure 1D) displays well-
resolved, sharp peaks communsurate with a kinetically
stable, interlocked species. Characteristic upfield shifts
of protons Hc, Hd, and He (0.92, 0.52, and 0.42 ppm,
respectively) are observed and are indicative18b,d,19g of
[π π] stacking interactions. An upfield shift of 0.52 ppm
3 3 3
is also observed for the aromatic proton Hb of the NDI
guest. The disappearance of maleimide proton Ha and the
formation of new signals in the 3.2ꢀ2.5 ppm region
indicate the formation of a thiol-maleimide Michael
adduct. Investigation of [2]rotaxane 5 by UV/vis spectro-
scopy reveals a charge-transfer band at 497 nm with
a molar extinction coefficient of ε = 714 Mꢀ1 cmꢀ1, which
is consistent18c,19e with other interlocked crown etherꢀ
naphthalene diimide systems.
(19) (a) Hamilton, D. G.; Sanders, J. K. M.; Davies, J. E.; Clegg, W.;
Teat, S. J. Chem. Commun. 1997, 897–898. (b) Try, A. C.; Harding,
M. H.; Hamilton, D. G.; Sanders, J. K. M. Chem. Commun. 1998, 723–
724. (c) Hamilton, D. G.; Davies, J. E.; Prodi, L.; Sanders, J. K. M.
Chem.;Eur. J. 1998, 4, 608–620. (d) Hansen, J. G.; Feeder, N.;
Hamilton, D. G; Gunter, M. J.; Becher, J.; Sanders, J. K. M. Org. Lett.
2000, 2, 449–452. (e) Hamilton, D. G.; Montalti, M.; Prodi, L.; Fontani,
M.; Zanello, P.; Sanders, J. K. M. Chem.;Eur. J. 2000, 6, 608–617. (f)
Gunter, M. J.; Bampos, N.; Johnstone, K. D.; Sanders, J. K. M. New J.
Chem. 2001, 25, 166–173. (g) Iijima, T.; Vignon, S. A.; Tseng, H.-R.;
Jarrosson, T.; Sanders, J. K. M.; Marchioni, F.; Venturi, M.; Apostoli,
E.; Balzani, V.; Stoddart, J. F. Chem.;Eur. J. 2004, 10, 6375–6392. (h)
Pascu, S. I.; Naumann, C.; Kaiser, G.; Bond, A. D.; Sanders, J. K. M.
Dalton Trans. 2007, 3864–3884.
One of the primary motivations of this work is the
development of efficient methods for integrating stimuli-
resonsive interlocked molecules with biological systems.
(21) (a) Asakawa, M.; Ashton, P. R.; Ballardini, R.; Balzani, V.;
Belohradsky, M.; Gandolfi, M. T.; Kocian, O.; Prodi, L.; Raymo, F. M.;
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Menzer, S.; Prodi, L.; Stoddart, J. F.; Venturi, M.; Williams, D. J.
Angew. Chem., Int. Ed. Engl. 1996, 35, 978–981.
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A.; Fyfe, M. C. T.; Gandolfi, M. T.; Gomez-Lopez, M.; Martınez-Dıaz,
´ ´
M.-V.; Piersanti, A.; Spencer, N.; Stoddart, J. F.; Venturi, M.; White,
A. J. P.; Williams, D. J. J. Am. Chem. Soc. 1998, 120, 11932–11942. (b)
Jones, J. W.; Gibson, H. W. J. Am. Chem. Soc. 2003, 125, 7001–7004. (c)
Ghosh, K.; Yang, H.-B.; Northrop, B. H.; Lyndon, M. M.; Zheng,
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Org. Lett., Vol. 14, No. 8, 2012

Scheme 2. Self-Assembly of N0-Boc-L-glutathione Dimethyl
Ester-Functionalized [2]Pseudorotaxane 7
As an initial, though important, first step toward this goal,
glutathione-functionalized thread 6 (Scheme 2) was
synthesized and its self-assembly with DNP38C10 was
investigated. A solvent mixture capable of solubilizing
both L-glutathione and macrocycle 3 could not be found;
therefore N0-Boc-L-glutathione dimethyl ester,23 which is
soluble in a range of organic solvents, was prepared.
Reacting this soluble glutathione derivative with thread 1
in CHCl3 and catalytic Et3N gave glutathione-functiona-
lized thread 6. Addition of 1.0 equiv of 3 to thread 6 in
CDCl3 resulted in a red solution indicating the formation
of [2]pseudorotaxane 7 (Scheme 2). [2]Pseudorotaxane 7
can also be obtained in one pot by adding catalytic
Et3N to a solution of 1:1 thread 1 and DNP34C10
macrocycle 3. UV/vis analysis of 7 (see Supporting
Information) reveals a CT band at 492 nm (ε =
137 Mꢀ1 cmꢀ1). Diagnostic protons of thread 7 and
macrocycle 3 were shifted upfield relative to their free
components as observed by 1H NMR spectroscopy
(Figure 2A). Shifts of 0.51, 0.96, 0.53, and 0.37 ppm
were observed for protons Hb, Hc, Hd, and He, respec-
tively. Notably, signals corresponding to both com-
plexed and uncomplexed species are well resolved in the
1H NMR spectrum obtained at 298 K and broadening
is essentially nonexistent at 268 K (Figure 2A). This
contrasts sharply with the significant line broadening
observed for [2]pseudorotaxane 4. It is hypothesized
that the bulkier glutathione moieties of thread 7 impose a
higher free energy barrier to threading/dethreading in
[2]pseudorotaxane 7 than the maleimide moieties of thread
1 in the case of [2]pseudorotaxane 4. Furthermore, the
greater kinetic stability of [2]pseudorotaxane 7 enabled
analysis by ESI/APCI mass spectrometry which indicated
a peak of m/z = 2305.88 [M þ Na]þ compared to a
calculated value of 2305.87 (Figure 2B). Peaks for both
Figure 2. (A) Partial 1H NMR spectrum (CDCl3, 268 K) of
glutathione-functionalized [2]pseudorotaxane 7. Protons are
labeled as in Scheme 1. Uncomplexed peaks are designated as
“(uc)”. (B) experimental (black solid line) and theoretical
(dashed red lines) ESI/APCI-MS isotopic distribution of 7.
macrocycle 3 and thread 6, however, could also be observed
in the same spectrum (see Supporting Information for the
full ESI/APCI-MS).
The results reported here highlight the utility of thiol-
maleimide click chemistry in the synthesis of mechanically
interlocked molecules. Some of the greatest prospects for
this chemistry stem from the potential to use thiol-maleimide
click chemistry as a means to incorporate mechanically
interlocked molecules into biological systems, opening the
possibility of using stimuli-responsive interlocked molecules
to impart allosteric control over the functions of proteins
and enzymes. The demonstration of glutathione-functiona-
lized [2]pseudorotaxane 7 represents a first step toward this
goal. Another key step to extending the chemistry presented
herein to the facile hybridization of interlocked molecules
with biological systems will be the synthesis of aqueous-
soluble derivatives of macrocycle 3. We are currently work-
ing toward these goals in order to develop the chemistry of
water-soluble proteinꢀrotaxane conjugates.
Acknowledgment. Acknowledgement is made to the
donors of The American Chemical Society Petroleum
Research Fund for partial support of this research.
Supporting Information Available. Synthetic proce-
1
dures, H and 13C NMR, mass spectroscopy, and UV/
vis spectroscopy. This material is available free of charge
via the Internet at http://pubs.acs.org.
(23) Flack, J. R.; Sangras, B.; Capdevila, J. H. Bioorg. Med. Chem.
2007, 15, 1062–1066.
The authors declare no competing financial interest.
Org. Lett., Vol. 14, No. 8, 2012
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