"Turn ON/OFF your LOV light": Borondipyrromethene-flavin dyads as biomimetic switches derived from the LOV domain

Article abstract of DOI:10.1002/anie.200462377

Molecular lamps: Analogously to a lamp with a light bulb, a switch, and a plug, the fluorescent ON state of dyad 1 can be blocked by other means besides the light switch ("removing the plug"). By using flavin as a redox unit and thermal or UV-activated th

Full text of DOI:10.1002/anie.200462377

Communications
Photoreceptor Mimetics
“Turn ON/OFF your LOV light”: Boron-
dipyrromethene–Flavin Dyads as Biomimetic
Switches Derived from the LOV Domain**
Christian Trieflinger, Knut Rurack,* and Jꢀrg Daub*
The induction and control of many important biological
processes rely on light signals. Chromophores participate in
actions such as charge separation, ion pumping, neural
responses, or photostimulated movement as photoreceptors
or photosensors.[1] In the course of evolution, Nature has
created rather complex biochemical processes that are photo-
regulated or -mediated, for example, the visual process or
light-activated plant defense. As light is a very convenient
tool to be used as a trigger or control signal, chemists have
been attracted by the potential that lies in photobiological
systems to be adopted for the design of artificial supramolec-
ular ensembles that show addressable photochemical activity.
Primary research areas here include photoresponsive mem-
branes, light-controlled host–guest chemistry, and photo-
chemical switching.[2] Among the latter, however, biologically
inspired systems are still scarce, and the examples reported so
far have been mainly accomplished in conjunction with
polymeric structures.[3]
Distinct molecular systems are even less abundant.[4] This
is surprising as particular biochemical moieties are very
attractive candidates for multimode supramolecular photo-
switches, a field of research in which we have become
increasingly interested over the years.[5] One such example is
the functionalized isoalloxazine chromophore. Flavins are
among the most important biological redox-active dye units,
and their rich photo, redox, and (inter)molecular chemistry
distinguish them as ideal composites for sophisticated chem-
ical switches.[6] Furthermore, a flavin residue is at the heart of
the active site of one of the most exciting photorecepting
proteins, the LOV (light-, oxygen-, or voltage-sensing)
[*] Dr. C. Trieflinger, Prof. J. Daub
Institut fꢀr Organische Chemie
Universitꢁt Regensburg
Universitꢁtsstrasse 31, 93053 Regensburg (Germany)
Fax: (+49)941-943-4984
E-mail: joerg.daub@chemie.uni-regensburg.de
Dr. K. Rurack
Div. I.3
Bundesanstalt fꢀr Materialforschung und -prꢀfung (BAM)
Richard-Willstꢁtter-Strasse 11, 12489 Berlin (Germany)
Fax: (+49)30-8104-5005
E-mail: knut.rurack@bam.de
[**] From the song by D. Malone and J. W. Scott (1961). LOV=light-,
oxygen-, or voltage-sensing domain (Science 1997, 278, 2120).
Financial support by the Deutsche Forschungsgemeinschaft (DFG)
and the Studienstiftung des Deutschen Volkes is highly appreciated.
This work is part of the Graduate College “Sensory Photoreceptors
in Natural and Artificial Systems” granted by DFG (GRK 640).
Supporting information for this article is available on the WWW
under http://www.angewandte.org or from the author.
2288
ꢀ 2005 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
DOI: 10.1002/anie.200462377
Angew. Chem. Int. Ed. 2005, 44, 2288 –2291

Angewandte
Chemie
domains of phototropin.[7,8] Based on this background and
owing to the exceptional complementarity of flavin and the
borondipyrromethene (BDP) fluorophore, which itself exhib-
its several important features of efficient photoswitching,[9]
we designed the BDP–flavin dyads 1–3 to realize our idea of a
distinctly controllable and reversible fluorescent switch (for
synthetic details, see Supporting Information).
phores, the main maxima being centered at 530 nm (BDP)
and 449 nm (flavin). This indication of electronically decou-
pled ground states is further supported by cyclic voltammetry,
in which the reversible reduction of flavin (À1.1 V vs. Fc+/Fc)
and the quasi-reversible oxidation and reduction of BDP
(0.65 V, À1.6 V vs. Fc+/Fc) are found at the expected
potentials. The different electrochemical behavior of 3
compared with 1 and 2, deducible from the additional
reoxidation peak at À0.92 V vs. Fc+/Fc (Table 1), is ascribed
to the well-known ECE (electron transfer–chemical step–
electron transfer) mechanism of flavin which involves an
intermolecular proton-assisted redox reaction.[10] The results
obtained for the ground-state properties of 1–3 presented so
far suggest that one important prerequisite for the facile
control of a switch is fulfilled, that is, both switching modules
are independent. However, to generate ON/OFF behavior as
expressed by strong signal changes in light output, a well-
defined OFF state is necessary, especially when a switch is
constructed from two strongly emitting fluorophores such as
flavin and BDP. In this respect, the suitability of our design
rationale is evident from Table 2 (see also the theoretical
considerations detailed in the Supporting Information).
Table 2: Fluorescence properties of 1–3.
Solvent
Ff ꢂ10À3 [a]
tf [ps][b]
tf [ns][c]
1
2
MeCN/toluene[d]
EtOH
1.3/2.3
1.6
0.7/1.3
1.0
0.8/1.5
8.8
10/17
13
6/11
8
6/10
96
–
5.33
–
5.54
–
–
MeCN/toluene[d]
EtOH
3
MeCN/toluene[d]
MeCN
3À [e]
[a] Relative to fluorescein 27 in NaOH (0.1n; 0.90Æ0.03),[15] Æ15%.
[b] At 298 K, Æ3 ps. [c] At 77 K, Æ0.003 ns. [d] First value in MeCN,
second in toluene. [e] Deprotonated with 1,8-diazabicyclo[5.4.0]undec-7-
ene (DBU).
The data in Table 1 and Figure 1 clearly reveal the
composite nature of the dyads as the absorption spectra of
1–3 are a linear combination of the bands of the subchromo-
Table 1: Absorption (in CHCl3) and redox data (in MeCN) of 1–3 (E1/2 in
All three dyads show only a weak fluorescence with
typical BDP features, regardless of solvent polarity and
proticity. No isoalloxazine emission could be detected, even
with excitation at 449 nm. The fluorescence also decays
rapidly, stressing the fact that an efficient nonradiative
deactivation path exists in the title compounds. Furthermore,
1 and 2 show a very similar behavior, excluding any influence
of a “meta” effect.[11] The important role of the directly fused
phenylene spacer becomes apparent, especially from the
emission data. This moiety acts simultaneously as an orbital
insulator (by preventing a mixing of states that is often found
in connection with the “meta” effect) and as an electronic
mediator (by enabling efficient fluorescence quenching). At
77 K, the fluorescence increases dramatically, and decay times
that are typical for BDP dyes and close to the value of the
reference compound 4 (6.27 ns) are measured. Again, after
excitation of the flavin, only BDP emission can be detected,
thus suggesting that efficient energy transfer to the BDP
occurs. At room temperature, photoinduced electron transfer
(PET) from the BDP to the electron-poor flavin most
probably accounts for the strong quenching of the fluores-
cence.
V vs. Fc+/Fc).
red
1=2
red
1=2
ox
1=2
labs [nm]
E
[V]
Ecp,red [V]
E
[V]
E
[V]
([lge]/[lg(mÀ1 cmÀ1)])
1
2
3
529 (4.86), 448 (4.15)
531 (4.81), 449 (4.12)
531 (4.81), 448 (4.10)
À1.11
À1.11
À1.10
–
–
À1.60
À1.63
À1.63
0.65
0.66
0.65
À0.92
Figure 1. Absorption spectra of 1 (solid line) and twofold reduced 1red
(c=5ꢂ10À6 m; dashed line) in MeCN/MeOH (1:1). Inset: Difference
spectrum of the flavin region.
Angew. Chem. Int. Ed. 2005, 44, 2288 –2291
www.angewandte.org
ꢀ 2005 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
2289

Communications
In glassy ethanol at 77 K, PET is inhibited as a result of the
mind the discourse in the introductory paragraph, at this stage
the photomediated reaction cycle of the LOV domain comes
into play. If a solution with the chemically activated ensemble
is irradiated with UV light of 254 nm, a strong increase in
BDP fluorescence is noticed. The high efficiency of the
switching process is strikingly evident from a several-100-fold
enhancement of BDP emission and is manifested in tf values
of 4.96 and 5.27 ns for the systems based on 1 and 2. The two
reduced dyads are fully and rapidly converted, and after
turning off the UV-light source, the fluorescence decreases
again to its initial value.
freezing out of solvent reorganization and intramolecular
movement. This interpretation is supported by the results
obtained upon deprotonation of 3 at N3. Such an increase in
electron density at the flavin entails a > 10-fold enhancement
of the BDP emission (Table 2). Dyads 1 and 2, which are
methylated at N3, do not show any spectroscopic changes
upon base addition.
Once the conventional OFF state of the switch was
defined, the next step was to install a control function that
allows this state to be “unlocked”. The versatile redox
chemistry of flavin[6] allows such an activation to be per-
formed chemically. In this experiment and in analogy to thiol-
based natural systems, the spectroscopic changes were
monitored in situ during the chemical reduction of the
flavin moiety with 1,3-propanedithiol[12] and DBU in
MeCN/MeOH (1:1) under an argon atmosphere. The prod-
ucts were the twofold reduced dyads 1red and 2red and
[1,2]dithiolane. As can be seen in Figure 1, the flavin
absorption at 440 nm decreases, whereas the BDP absorption
is not affected, which clearly indicates that the redox process
occurs in the flavin. However, both 1red and 2red are still
weakly fluorescent, with fluorescence lifetimes within the
resolution of the instrument (tf = 3–5 ps). These observations
now mark the chemically controlled OFF state and can be
explained by a reversal of the ET process (Supporting
Information). In the reduced dyads, the electron density at
the flavin is so high that excitation of the BDP triggers a PET
from the flavin to the BDP, reductive PET processes being
well-known for donor-substituted BDPs (Figure 2).[9b]
The reversibility of this reaction is shown in Figure 2,
which indicates that irreversible side reactions do not play a
major role. Most probably, UV irradiation initiates the
formation of a thermally unstable but highly emissive species,
which is accumulated upon continuous irradiation. Dyads 1red
and 2red are then regenerated in a thermal reverse reaction.
Although the real nature of the emitting species could not yet
be analytically determined with methods available to us, the
results strongly suggest that the electron density of the flavin
is considerably lower than in the oxidized state, but higher
than in the fully reduced state. A mono-reduced state can be
excluded as such radicals would also quench the emission.
Thus, we tentatively assume that the light-induced for-
mation of an emissive flavin–thiol adduct in analogy to the
LOV domain photocycle occurs under the prevailing reaction
conditions. The thermal recovery rates reported for LOV
domains are commonly in the range of 0.02 to 0.003 sÀ1,
depending on its type, the medium, and possible protein
assistance.[7,13] These properties suggest that a C4a adduct or a
C4a–C10a cycloadduct might be involved under the present,
more-extreme conditions. Further experiments are currently
underway to elucidate the mechanism in more detail.
In conclusion we have presented novel BDP–flavin dyads
with distinctive electrochemical and optical properties,
including energy- and electron-transfer-modulated emission.
The direction and efficiency of the ET process can be
influenced by various input signals, such as the addition of
base, redox processes, and irradiation with light. These
features distinguish 1 and 2 as potent biomimetic photo-
chemical switches[14] in which the activation state can be
independently controlled and the output emission reversibly
switched.
Now that we were able to switch between an inactive and
an activated OFF state, the last requirement concerned the
input that allows reversible OFF/ON switching. Keeping in
Received: October 20, 2004
Published online: March 8, 2005
Keywords: boron · flavin · fluorescence · molecular switches ·
.
photoreceptors
[1] M. A. van der Horst, K. J. Hellingwerf, Acc. Chem. Res. 2004, 37,
13 – 20.
Figure 2. Reaction scheme of the thermal and photochemical switch-
ing between the non-activated OFF states (1, 2), the activated OFF
states (1red, 2red), and the ON state (reporter state). Inset: Reversible
switching between the activated OFF (mauve) and the ON state
(yellow) of 1red as reflected by the change in emission intensity in
MeCN/MeOH (1:1) during several irradiation cycles with cycle times
of 30 s irradiation and 300 s thermal reverse reaction (c1 =1ꢂ10À6 m).
[2] a) T. Kinoshita, J. Photochem. Photobiol. B 1998, 42, 12 – 19;
b) I. Willner, B. Willner in Molecular Switches (Ed.: B. L.
Feringa), Wiley-VCH, Weinheim, 2001, pp. 165 – 218.
[3] a) O. Pieroni, A. Fissi, N. Angelini, F. Lenci, Acc. Chem. Res.
2001, 34, 9 – 17; b) I. Willner, S. Rubin, React. Polym. 1993, 21,
177 – 186.
2290
ꢀ 2005 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.angewandte.org
Angew. Chem. Int. Ed. 2005, 44, 2288 –2291

Angewandte
Chemie
[4] a) A. Niemz, V. M. Rotello, Acc. Chem. Res. 1999, 32, 44 – 52;
b) Z. Shen, R. Prochazka, J. Daub, N. Fritz, N. Acar, S.
Schneider, Phys. Chem. Chem. Phys. 2003, 5, 3257 – 3269.
[5] a) H. Spreitzer, J. Daub, Chem. Eur. J. 1996, 2, 1150 – 1158; b) K.
Rurack, M. Kollmannsberger, J. Daub, Angew. Chem. 2001, 113,
396 – 399; Angew. Chem. Int. Ed. 2001, 40, 385 – 387; c) K.
Rurack, A. Kovalꢀchuck, J. L. Bricks, J. L. Slominskii, J. Am.
Chem. Soc. 2001, 123, 6205 – 6206.
[6] Chemistry and Biochemistry of Flavoenzymes, Vol. 1 (Ed.: F.
Mꢁller), CRC, Boca Raton, FL, 1991.
[7] a) T. E. Swartz, S. B. Corchnoy, J. M. Christie, J. W. Lewis, I.
Szundi, W. R. Briggs, R. A. Bogomolni, J. Biol. Chem. 2001, 276,
36493 – 36500; b) E. Schleicher, R. M. Kowalczyk, C. W. M.
Kay, P. Hegemann, A. Bacher, M. Fischer, R. Bittl, G. Richter, S.
Weber, J. Am. Chem. Soc. 2004, 126, 11067 – 11076.
[8] LOV domains are the primary signaling subunits of blue-light
photoreceptors and sense light, oxygen, or voltage. They transfer
the information to serine/threonine kinases. In the primary step,
a C4a-thiol adduct of flavin mononucleotide is formed.
[9] a) R. A. Wagner, J. S. Lindsey, J. Am. Chem. Soc. 1994, 116,
9759 – 9760; b) M. Kollmannsberger, T. Gareis, S. Heinl, J. Breu,
J. Daub, Angew. Chem. 1997, 109, 1391 – 1393; Angew. Chem.
Int. Ed. Engl. 1997, 36, 1333 – 1335; c) M. Kollmannsberger, K.
Rurack, U. Resch-Genger, W. Rettig, J. Daub, Chem. Phys. Lett.
2000, 329, 363 – 369.
[10] A. Niemz, J. Imbriglio, V. M. Rotello, J. Am. Chem. Soc. 1997,
119, 887 – 892.
[11] F. D. Lewis, J.-S. Yang, J. Am. Chem. Soc. 1997, 119, 3834 – 3835.
[12] E. L. Loechler, T. C. Hollocher, J. Am. Chem. Soc. 1980, 102,
7312 – 7321.
[13] M. Salomon, J. M. Christie, E. Knieb, U. Lempert, W. R. Briggs,
Biochemistry 2000, 39, 9401 – 9410.
[14] For a native LOV domain as a photochromic switch, see: J. T. M.
Kennis, I. H. M. van Stokkum, S. Crosson, M. Gauden, K.
Moffat, R. van Grondelle, J. Am. Chem. Soc. 2004, 126, 4512 –
4513.
[15] J. Olmsted III, J. Phys. Chem. 1979, 83, 2581 – 2584.
Angew. Chem. Int. Ed. 2005, 44, 2288 –2291
www.angewandte.org
ꢀ 2005 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
2291