Fluorescence resonance energy transfer across a mechanical bond of a rotaxane

Article abstract of DOI:10.1039/b506177f

Fluorescence transfer across a donor-acceptor tagged rotaxane was studied and a small conformational change of the rotaxane observed using fluorescent spectroscopy and ROESY NMR. The Royal Society of Chemistry 2005.

Full text of DOI:10.1039/b506177f

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Fluorescence resonance energy transfer across a mechanical bond of a
rotaxane{
Hideki Onagi and Julius Rebek, Jr.*
Received (in Columbia, MO, USA) 3rd May 2005, Accepted 6th July 2005
First published as an Advance Article on the web 18th August 2005
DOI: 10.1039/b506177f
343 was the acceptor on the macrocycle. The motion of the
interlocked molecule was based on the behavior of rotaxanes
developed by Leigh and co-workers12 (Fig. 1b). The most
straightforward synthesis, involving macrocyclization13 of the fully
labeled components around 2 was thwarted by low solubility.
Instead, an extended alkyl chain was attached to the macrocycle
precursor 3 on an Alloc group using olefin metathesis (see
electronic supplementary information). The macrocycliation of
precursor 3 with 4 to gave the [2]rotaxane 5 (Scheme 1). The
standard deprotection using Pd(0) removed the modified
Alloc group, and gave the amino-[2]rotaxane 6. The installation
of coumarin 343 on the macrocycle through the amide condensa-
tion using EDCI concluded the synthesis of the FRET labeled
[2]rotaxane 1.
Fluorescence transfer across a donor-acceptor tagged rotaxane
was studied and a small conformational change of the rotaxane
observed using fluorescent spectroscopy and ROESY NMR.
Fluorescence resonance energy transfer (FRET) is widely used
to monitor asssociation and dynamic processes of biological
molecules,1 and has been used to monitor the self-assembly of
molecular capsules.2 There are only a few systems featuring
FRET across mechanically interlocked components,3–7 and
we proposed a new rotaxane with FRET labels. Rotaxanes
have been extensively studied for the development of mechanically
bonded molecular machines8 (Fig. 1a),8d,e such as shuttles,9
switches10 and even muscles.11 We report here the synthesis and
characterization of the system, and observations relevant to their
use as moving parts.
The absorption and emission spectra of the [2]rotaxane 1
were recorded as illustrated in Fig. 2. Labeled 1 possesses the
characteristic absorption of both donor and acceptor coumarins,
and the fluorescence of the acceptor (lmax 5 470 nm in CHCl3)
when excited at 345 nm shows that efficient FRET occured. This
emission property of 1 was not affected under high dilution, as
The relevant structural feature of the rotaxane 1 is a macrocycle
bearing a fluorophore and the axle bearing another fluorophore
at one end. Coumarin 2 was the donor on the axle and coumarin
Fig. 1 (a) Schematic representation of the rotaxane showing that a
macrocycle (red) was mechanically interlocked by a dumbbell composed of
an axle (black) with a hydrogen bonding site (green), terminated by the
bulky end group (yellow). Interlocked components were tagged by
the donor D (blue) and acceptor A (purple). (b) Chemical structure of the
[2]rotaxane 1.
The Skaggs Institute for Chemical Biology and The Department of
Chemistry, The Scripps Research Institute, MB-26, 10550 North Torrey
Pines Road, La Jolla, CA 92037, USA. E-mail: jrebek@scripps.edu;
Fax: +1 858-784-2876; Tel: +1 858-784-2250
{ Electronic supplementary information (ESI) available: Synthetic and
characterisation details for 1 and additional fluorescence experiments. See
http://dx.doi.org/10.1039/b506177f
Scheme 1 Synthesis of the [2]rotaxane 1: (i) Et3N, anhydrous CHCl3,
RT, 3 and 4 were added to a solution of 2 by syringe pump over 4 h then
stirred for 10h; (ii) Pd(PPh3), PhSiH3, CH2Cl2, RT, 12 h, 8.4% (through
procedures i and ii); (iii) EDCI, CHCl3, RT, 10h, 69%.
4604 | Chem. Commun., 2005, 4604–4606
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Although it is clear that the overall intensity of both absorption
and emission are lower in polar solvent (DMSO), the fluorescence
intensity in DMSO is relatively lower than that in CHCl3. This
can be attributed to the release of the macrocycle from the
Gly-Gly station through the disruption of hydrogen bonding
by DMSO. The Fo¨rster distances of the dyes are typically around
1a
20–80 A; a distance that mostly exceeds the length of the axle.
˚
Accordingly, the system maintains FRET as long as the rotaxane
is intact.
The motion of the macrocycle was also observed using NMR
analysis. The conformation of the rotaxane 1 was elucidated by 1H
and ROESY NMR spectra as shown in Fig. 3. In a non-polar
environment,14 the macrocycle was predominantly located around
one of the glycine residues and the proton Ha. In the polar
environment, the macrocycle shifts towards the center of the axle
as is apparent from the upfield shifts (up to 0.8 ppm) of protons
that also show NOE interactions with the macrocycle.
In summary, the present research demonstrated that FRET
labeling can be used to detect small motions in mechanically
interlocked systems.
Fig. 2 Absorption (—) and fluorescence (......) spectra of the [2]rotaxane
1 in CHCl3 (red) and DMSO (blue), (50 mM, lexc 5 345 nm, 298 K).
expected by the mechanical bonding of the donor and acceptor
(see ESI{). Absorption and fluorescence properties of 1 were
also investigated under polar and non-polar environments.
We are grateful to the Skaggs Institute for Research and to The
National Institutes of Health (GM 50174) for financial support.
H. O. is a Skaggs Postdoctoral Fellow.
Notes and references
1 For reviews on FRET theory and applications, see: (a) P. Wu and
L. Brand, Anal. Biochem., 1994, 218, 1; (b) J. Szo¨llo˝si, S. Damjanovich
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2 R. K. Castellano, S. L. Craig, C. Nuckolls and J. Rebek, Jr., J. Am.
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3 M. Tamura and A. Ueno, Chem. Lett., 1998, 369.
4 For related examples where a macrocycle itself quenches fluorescence of
a donor, see: (a) G. Giro, M. Cocchi, V. Fattori, G. Gadret, G. Ruani,
M. Cavallini, F. Biscarini, R. Zamboni, T. Loontjens, J. Thies,
D. A. Leigh, A. F. Morales and R. F. Mahrt, Synth. Met., 2001, 122,
63; (b) E. M. Perez, D. T. F. Dryden, D. A. Leigh, G. Teobaldi and
F. Zerbetto, J. Am. Chem. Soc., 2004, 126, 12210.
5 For related systems but fluorophores are covalently attached, see: (a)
D. H. Qu, G. C. Wang, J. Ren and H. Tian, Org. Lett., 2004, 6, 2085;
(b) K. Hiratani, M. Kaneyama, Y. Nagawa, E. Koyama and
M. Kanesato, J. Am. Chem. Soc., 2004, 126, 13568.
6 For examples of non-interlocked systems, see: (a) E. Ishow, A. Credi,
V. Balzani, F. Spadola and L. Mandolini, Chem. – Eur. J., 1999, 5, 984;
(b) L. Jullien, J. Canceill, B. Valeur, E. Bardez, J.-P. Lefe`vre, J.-M. Lehn,
V. Marchi-Artzner and R. Pensu, J. Am. Chem. Soc., 1996, 118, 5432.
7 For examples of electron/energy transfer, see: (a) M. Andersson,
M. Linke, J.-C. Chambron, J. Davidsson, V. Heitz, L. Hammarstro¨m
and J.-P. Sauvage, J. Am. Chem. Soc., 2002, 124, 4347; (b) K. Li,
´
P. J. Bracher, D. M. Guldi, M. A. Herranz, L. Echegoyen and
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8 For reviews on molecular machines, see: (a) P. F. Barbara, Acc. Chem.
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J. F. Stoddart; (b) V. Balzani, M. Venturi and A. Credi, Molecular
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Weinheim, 2003; (c) C. J. Easton, S. F. Lincoln, L. Barr and H. Onagi,
Chem.-Eur. J., 2004, 10, 3120. Specific to rotaxanes and catenanes, see:
(d) V. Balzani, M. Go´mez-Lo´pez and J. F. Stoddart, Acc. Chem. Res.,
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1
1
Fig. 3 A section of the 600 MHz H spectra of the axle 2 and H and
ROESY NMR spectra of the [2]rotaxane 1 recorded in CDCl3 + CD3OD
(600 : 50 mL) and d6-DMSO, showing the NOEs between the axle (x-axis)
and the HA–A9 protons of the macrocycle (y-axis).
10 (a) M. Cavallini, F. Biscarini, S. Leon, F. Zerbetto, G. Bottari and
D. A. Leigh, Science, 2003, 299, 531; (b) A. H. Flood, J. F. Stoddart,
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11 M. C. Jime´nez, C. Dietrich-Buchecker and J.-P. Sauvage, Angew.
Chem., Int. Ed., 2000, 39, 3284.
12 For example, see: T. Da Ros, D. M. Guldi, A. F. Morales, D. A. Leigh,
M. Prato and R. Turco, Org. Lett., 2003, 5, 689.
13 A. G. Johnston, D. A. Leigh, R. J. Pritchard and M. D. Deegan,
Angew. Chem., Int. Ed., 1995, 34, 1209.
14 In order to determine the conformation in a non-polar environ-
ment, a mixture of CDCl3 with a small amount of CD3OD
was used. This is because the spectrum of 1 in pure CDCl3 was
broad (258 K to 308 K), and was not suitable for accurate
analysis. The broadening is caused by slow rotation of the macrocycle
on the axle.
4606 | Chem. Commun., 2005, 4604–4606
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