Adhesive Photoswitch: Selective Photochemical Modulation of

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Adhesive Photoswitch: Selective Photochemical Modulation of Enzymes under Physiological Conditions Rina Mogaki,† Kou Okuro,*,† and Takuzo Aida*,†,‡ †

Department of Chemistry and Biotechnology, School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8656, Japan ‡ Riken Center for Emergent Matter Science, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan S Supporting Information *

ABSTRACT: We developed a water-soluble adhesive photoswitch (Gluen-Azo-SA, average n = 5) that selectively binds to a target enzyme and photochemically modulates its enzymatic activity even in cell lysates. Its design strategy features a photochromic azobenzene unit (Azo), which carries on one side an inhibitory motif for the target enzyme and on the other a glue part (Gluen) that utilizes its multiple guanidinium ion (Gu+) pendants for adhering onto the target surface. The target of Gluen-Azo-SA is carbonic anhydrase (CA) because sulfonamide (SA) derivatives, such as SA at the terminus of Gluen-Azo-SA, are known to bind selectively to the CA active site. The SA moiety, upon docking at the CA active site, possibly guides the connecting Gluen part to an oxyanion-rich area in proximity to the active site for adhesion, so that the conjugation between Gluen-Azo-SA and CA is ensured. With this geometry, the photochemical isomerization of the Azo unit likely generates a push−pull motion of SA, resulting in its docking and undocking at the active site of CA with the help of a competing substrate. Consequently, Gluen-Azo-SA can selectively photomodulate the enzymatic action of CA even in cell lysates. Azo-SA without Gluen can likewise suppress the enzymatic activity of CA by docking SA at its active site, but the resulting CA/Azo-SA conjugate, in contrast, does not respond to light.

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INTRODUCTION Photochemical modulation of biomolecular functions may provide noninvasive methods to cure diseases and is also interesting for elucidating biological events.1 A promising approach along this line features site-specific anchoring of a “photoswitch” onto a target protein. In general, a “photoswitch” consists of a photoisomerizable unit in conjunction with an inhibitor or activator.2 It should adopt a particular geometry such that one of its isomeric forms is more favorable than the other for proper localization of the inhibitor or activator moiety onto the guest-binding site.2a,e Consequently, one can photochemically modulate certain biological functions of proteins.2 Based on the strategy described above, many photoswitchable functional proteins have been successfully developed. In 1971, Erlanger and co-workers first reported a photoswitchable nicotinic acetylcholine receptor.3 In the past decade, Isacoff, Kramer, Trauner, and co-workers generalized this concept4−6 and contributed significantly to the development of a variety of photomodulable ion channels, such as potassium ion channels4 and ionotropic glutamate receptors (iGluR).5 Furthermore, photomodulable iGluR was applied to the control of zebrafish behaviors5b,d and restoration of the light sensitivity in blind mice.5e,f Recently, Chin and co-workers achieved incorporation of a photoswitch in living cells using a bio-orthogonal Diels− Alder reaction with genetically engineered MAPK/ERK kinase © 2017 American Chemical Society

(MEK) containing an unnatural amino acid unit for conjugation.7 One important issue is that the “photoswitches” so far reported often require genetic mutation of the target proteins for covalent anchoring of the switching moiety.4a,b,5,6b This approach is appropriate for realizing a high spatial resolution. However, in view of the practicality, noncovalent “photoswitches”, which do not require covalent anchoring, may be preferred. Previously, several types of noncovalent photoswitches have been developed, particularly for cell membrane proteins such as neurotransmitter receptors8 and ion channels.9 A representative example is azobenzene derivatives of propofol developed by Trauner and co-workers, which allowed photoswitching of the signal transduction of a γ-aminobutyric acid (GABA) receptor in living tadpoles.8c They also achieved photochemical modulation of tubulin polymerization dynamics in living cells using azobenzene derivatives of combretastatin A-4 as photoswitchable inhibitors.10 In addition to the receptor and ion channel proteins, enzymes are one of the most attractive targets for photochemical modulation since they play important roles in cell homeostasis and functions. Up to date, photoisomerizable inhibitors for intracellular enzymes such as DNA Received: May 18, 2017 Published: July 4, 2017 10072

DOI: 10.1021/jacs.7b05151 J. Am. Chem. Soc. 2017, 139, 10072−10078

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Journal of the American Chemical Society

Figure 1. Schematic structures of (a) adhesive photoswitch Gluen-Azo-SA (average n = 5) consisting of adhesive Gluen, photochromic Azo, and enzymatic inhibitor sulfonamide (SA) for CA, (b) Gluen-Azo (average n = 4), a reference without inhibitory SA, (c) Gluen-Cy5.5 (average n = 5) consisting of adhesive Gluen and fluorescent dye Cy5.5, (d) Azo-SA, a reference without adhesive Gluen, and (e) sulfanilamide, a noncovalent alternative to the SA moiety in Gluen-Azo-SA. (f) Schematic representation of the trans-to-cis and cis-to-trans isomerizations of the Azo moiety in response to the irradiation with UV and visible light, respectively.

gyrase,11a acetylcholinesterase,11b proteasome,11c and histone deacetylase11d,e have been studied, and it was reported that their isomers exhibit different inhibitory effects from one another. However, in situ photochemical switching between different enzymatic activities in intracellular environments still remains a big challenge. In the present work, we developed a water-soluble adhesive photoswitch (Gluen-Azo-RInhibit) that selectively binds to a target enzyme and photochemically modulates its enzymatic activity even in cell lysates as a model of intracellular environments. The design strategy of Gluen-Azo-RInhibit features a photochromic azobenzene unit (Azo), which carries on one side an inhibitory motif (RInhibit) for a target enzyme and on the other a glue part (Gluen) that utilizes its multiple guanidinium ion (Gu+) pendants for adhering onto the target enzyme. As a proof-of-concept study, we utilized for RInhibit a sulfonamide (SA, Figure 1a; Gluen-Azo-SA, average n = 5) derivative, which is known as a carbonic anhydrase (CA) inhibitor that binds to its active site.12 As the adhesive motif, we took note of molecular glues.13,14 Previously, we developed water-soluble molecular glues13,14 bearing multiple guanidinium ion (Gu+) pendants,15 which tightly adhere to proteins,13a,c,e,f,14b,e nucleic acids,14c,d phospholipid membranes,13d and clay nanosheets13b,14a via the formation of multiple salt bridges between their Gu+ pendants and oxyanionic groups on the targets.15a The conjugation of a molecular glue to an enzyme inhibitor increases the binding affinity due to the salt-bridge-based immobilization of the inhibitor to the protein surface; thereby, an enhanced inhibitory effect can be expected.14b We envisioned that, by docking to the active site of CA, SA could properly guide the connecting Gluen part to an oxyanionrich area on CA in proximity to its active site for a salt-bridge interaction (Figure 2a),16 so that the conjugation between

Gluen-Azo-SA and CA is ensured.17 With this geometry, the photochemical isomerization of the Azo unit (Figure 1f) likely generates a push−pull motion of SA, resulting in its docking and undocking at the active site of CA (Figure 2c) with the help of a competing substrate if present (Figure 2b). Our results unprecedentedly demonstrated that Glue5-Azo-SA (Gluen-Azo-SA, average n = 5; Figure 1a) enables selective photochemical modulation of the enzymatic activity of CA (Figure 2c) even under physiological conditions such as in cell lysates that consist of intracellular components.

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RESULTS AND DISCUSSION Glue5-Azo-SA (Figure 1a) was synthesized using a “click” reaction18,19 between Azo-SA (Figure 1d) and a four-armed monomer containing azide, alkyne, and two Boc-protected Gu+ pendants, followed by the removal of the Boc groups. A reference polymer without SA (Gluen-Azo; average n = 4; Figure 1b) was likewise synthesized using N3-appended Azo, instead of Azo-SA. Fluorescently labeled Gluen-Cy5.5 (average n = 5; Figure 1c) was also synthesized using N3-Cy5.5. Average numbers of the repeating units (n) in Gluen-Azo-SA, Gluen-Azo, and Gluen-Cy5.5 were evaluated by 1H NMR end-group analysis. Prior to the enzymatic activity assays, we investigated the photoisomerization kinetics of Glue5-Azo-SA by electronic absorption spectroscopy. When a HEPES buffer (50 mM, pH 7.2) solution of Glue5-Azo-SA (10 μM) at 37 °C was exposed to UV light at 365 nm, the absorbance at 360 nm, due to the trans form of the Azo unit, decreased with an increase in the absorbance at 440 nm due to the cis form (Figure 3a). These spectral changes leveled off after irradiation for 24 s due to the establishment of the photostationary state. Subsequently, photoirradiation with visible light (>400 nm) for 2 s resulted 10073


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Figure 2. (a) Schematic illustration of how adhesive photoswitch Gluen-Azo-SA (RInhibit = sulfonamide SA) interacts with carbonic anhydrase (CA)16 as a target enzyme. Ideally, the SA moiety, with a higher binding affinity toward CA than the Gluen part (the association constants: KGlue < KInhibit), primarily determines a proper location of Gluen-Azo-SA on CA by docking at its active site and then guides the Gluen part to an oxyanion-rich proximal area to adhere to, thereby ensuring the CA/Gluen-Azo-SA conjugation. (b) In the actual catalytic system involving a large amount of substrate p-nitrophenyl acetate (PNPA), the binding of the SA moiety to the active site of CA occurs competitively with PNPA, so that the KInhibit value is considerably decreased to such a level that is comparable to that of the Gluen part (KGlue). (c) In such a case, the geometrical change of the Azo unit in Gluen-Azo-SA by irradiation with UV and visible light can be converted into a mechanical push−pull motion of the SA moiety, resulting in its docking and undocking at the active site of CA. Hence, the CA/Gluen-Azo-SA conjugate photochemically increases and decreases its hydrolytic activity. (d) If Azo-SA has no adhesive part (Gluen), the isomerization effect of the Azo unit on the binding affinity of SA would be negligibly small.

states upon exposure to UV (365 nm) and visible (425 nm) light, respectively (Figure S12).20 Gluen adheres to the CA surface, as expected from the distribution of its negative charges (Figure S19).20 When CA, fluorescently labeled with Cy5 (Cy5CA; 1 μM),20 was titrated with Glue5-Cy5.5 (0−60 μM) at 37 °C in HEPES buffer (50 mM, pH 7.2), the fluorescence at 656 nm (λex = 639 nm), attributable to the emission of Cy5, was partially quenched (Figure 4a), indicating the occurrence of a fluorescence resonance energy transfer (FRET) from Cy5 to Cy5.5.21 According to a previously published method,22 we fitted the observed fluorescence intensity change (Figure 4b) to a 1:1 binding model and obtained an association constant (Kassoc) of Glue5-Cy5.5 toward Cy5CA as 2.6 ± 1.4 × 105 M−1. Dansylamide (DNSA) is known to be bound to the active site of CA23 with a Kassoc value, as obtained in the present study, of 2.6 ± 0.4 × 106 M−1 at 25 °C in HEPES buffer (50 mM HEPES, pH 7.2; Figure S14).20 DNSA, when bound to the active site of CA, fluoresces at 470 nm (λex = 280 nm; Figure 4c) due to a FRET from the photoexcited tryptophan residues near the active site.23 We found that the addition of Glue5-AzoSA (0−5 μM) to a CA (0.25 μM)/DNSA (5 μM) conjugate at 25 °C in HEPES buffer results in a decrease in fluorescence intensity at 470 nm (Figure 4d). Meanwhile, such a fluorescence quenching feature was not observed when Glue4-Azo (0−5 μM) without the SA moiety was used under

Figure 3. Absorption spectral changes of Glue5-Azo-SA (10 μM) at 37 °C in HEPES buffer (50 mM, pH 7.2, 600 μL) upon (a) photoirradiation at 365 nm (0−24 s, 100 W) and then (b) at >400 nm (0−2.0 s, 100 W).

in an opposite spectral change to the above (Figure 3b), where the absorbance at 360 nm increased at the expense of the absorbance at 440 nm. Accordingly, the spectral change profile was virtually identical to that of Azo-SA without Gluen (Figure S8),20 indicating that the photoisomerization behavior of the Azo unit is intact to the conjugation with Gluen. The half-life of cis-Glue5-Azo-SA at 37 °C in HEPES buffer was 2.3 h in the dark (Figure S11b).20 This slow thermal backward isomerization is advantageous for producing a large steady state fraction of the cis form. As a reference, the estimated trans/cis ratios of Azo-SA in DMSO-d6 by means of 1H NMR spectroscopy were 7/93 and 76/24 at the photostationary 10074


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noncovalent inhibitors. CA catalyzes the interconversion between H2O/CO2 and H+/HCO3− and plays an essential role in many physiological processes, including gas exchange and pH homeostasis.26 CA also exhibits an esterase activity.27 Therefore, when p-nitrophenyl acetate (PNPA, 1.2 mM) as a substrate was mixed with CA (1 μM) at 37 °C in HEPES buffer (50 mM, pH 7.2), PNPA was hydrolyzed to generate pnitrophenol (Figure 5a), resulting in an increase in its

Figure 4. (a) Fluorescence spectral change of Cy5-labeled CA (Cy5CA, 1 μM; λex = 639 nm) at 37 °C in HEPES buffer (50 mM, pH 7.2) upon titration with Glue5-Cy5.5 (0−60 μM) and (b) its titration profile at 656 nm. The association and dissociation constants (Kassoc and Kd, respectively) represent the means ± SD. (c) Schematic illustration of the competitive binding of Glue5-Azo-SA with dansylamide (DNSA) to the active site of CA. DNSA, which fluoresces upon binding to the active site of CA, is not fluorescent when kicked out from the active site.23 (d) Fluorescence spectral change (λex = 280 nm) of a mixture of CA (0.25 μM) and DNSA (5 μM) at 25 °C in HEPES buffer (50 mM, pH 7.2) upon titration with Glue5-Azo-SA (0−5 μM) and (e) the dissociation profile of DNSA from the active site of CA. The fractions of DNSA-bound CA were calculated from the fluorescence intensity at 470 nm.20 The concentrations of free Glue5-Azo-SA were calculated from the fractions of DNSA-bound CA. The Kassoc and Kd values represent the means ± SD.

Figure 5. (a) CA-mediated enzymatic hydrolysis of p-nitrophenyl acetate (PNPA) to p-nitrophenol, exhibiting its characteristic absorption at 405 nm.27 (b) Absorption spectral changes at 405 nm of a mixture of PNPA (1.2 mM) and CA (1 μM) in HEPES buffer (50 mM, pH 7.2) at 37 °C in the absence (black) and presence of Glue5Azo-SA (2.5 μM; red), Azo-SA (2.5 μM; blue), Glue4-Azo (2.5 μM; green), and a mixture of Glue4-Azo (2.5 μM) and sulfanilamide (2.5 μM; orange). The sample solutions were exposed to UV (365 nm, 20 s; purple-shaded areas) and visible light (>400 nm, 5 s; pink-shaded areas) alternately. (c) Absorption spectral changes at 405 nm of a mixture of PNPA (1.2 mM) and CA (1 μM) in Hep3B cell lysates (9.0 × 103 cell equivalents/mL, pH 7.2)20 at 37 °C in the absence (black) and presence of Glue5-Azo-SA (3 μM; red). The sample solutions were exposed to UV (365 nm, 20 s; purple-shaded areas) and visible light (>400 nm, 5 s; pink-shaded areas) alternately.

conditions otherwise identical to the above (Figure S16).20 These results indicated that DNSA at the active site of CA was kicked out by the SA moiety of Glue5-Azo-SA (Figure 4c). By fitting the dissociation profile of DNSA (Figure 4e), as evaluated from the decrease in fluorescence intensity at 470 nm,20 to the competitive binding model,24 the Kassoc value of Glue5-Azo-SA toward the active site of CA was estimated as 9.3 ± 3.5 × 107 M−1, which is significantly larger than that observed for Glue5-Cy5.5/Cy5CA (Figure 4b; Kassoc = 2.6 ± 1.4 × 105 M−1). Azo-SA also exhibited a larger Kassoc value toward CA (6.8 ± 3.0 × 107 M−1; Figure S15)20 than the Glue5 part. Thus, the binding of the SA moiety of Glue5-Azo-SA to the active site of CA would occur more preferentially than the adhesion of the Glue5 part (Figure 2a). We then investigated the possibility of in situ photoswitching of the enzymatic activity of CA by the action of Glue5-Azo-SA. Previously, SA-appended noncovalent photoisomerizable inhibitors for CA were reported by using dithienylethene as a photochromic unit.25 However, in situ photoswitching of the enzymatic activity of CA has not been reported with such

characteristic absorption at 405 nm (Figure 5b, black, 0−120 s; ΔAbs/s = 2.0 × 10−3).27 When treated with Glue5-Azo-SA (2.5 μM), CA decreased its hydrolytic activity to 23% (Figure 5b, red, 0−240 s; ΔAbs/s = 4.5 × 10−4). Subsequent 20 s UV exposure at 365 nm resulted in the nearly complete recovery of the intrinsic activity of CA (Figure 5b, red, 240−360 s; ΔAbs/s = 1.9 × 10−3). When the reaction mixture was then exposed to visible light for 5 s (>400 nm), the hydrolytic reaction again 10075


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Journal of the American Chemical Society decreased to 25% (Figure 5b, red, 360−480 s; ΔAbs/s = 4.9 × 10−4). As shown in Figures 5b and 6 (red),28 the CA/Glue5-

tion of the Azo unit, while the SA moiety remains at the active site of CA. In regard to this issue, it should be noted that SA is a competitive inhibitor for CA. So, in the presence of substrate PNPA (1.2 mM), the Kassoc value of the SA moiety to the active site of CA (1 μM) is roughly two orders of magnitude smaller (9.3 ± 3.5 × 107 M−1 → 8.9 ± 0.5 × 105 M−1; Table 1 and Figure S17a),20 which is nearly comparable to that of the Glue5 part toward CA (Figure 4b). Further to note, when the reaction was carried out after 1 min UV exposure under conditions otherwise identical to the above, the Kassoc value of Glue5-AzoSA observed was 5.2-fold smaller (Kassoc = 1.7 ± 0.1 × 105 M−1) than that before the UV exposure (Table 1 and Figure S17a).20 Hence, in the actual catalytic system, the photoisomerization of the Azo unit in Glue5-Azo-SA may possibly result in the preferential detachment of the SA moiety from the active site of CA, leading to the photomodulation of the hydrolytic process of PNPA (Figure 2c). As a reference, Azo-SA without Glue5 barely changed its Kassoc value upon 1 min UV exposure (4.0 ± 2.1 × 105 M−1 → 3.1 ± 0.7 × 105 M−1; Table 1 and Figure S17b).20 Finally, we investigated the CA (1 μM)-mediated hydrolysis of PNPA (1.2 mM) at 37 °C in the lysates of human hepatocellular carcinoma Hep3B cells (9.0 × 103 cell equivalents/mL, pH 7.2),20 as a mimic of the intracellular media, in order to investigate how Glue5-Azo-SA (3 μM) operates for CA in the coexistence of a variety of competing oxyanionic biomolecular species. In the cell lysates without Glue5-Azo-SA, PNPA was hydrolyzed faster (Figure 5c, black) than that in HEPES buffer (Figure 5b, black), possibly due to the action of esterases derived from Hep3B cells. Noteworthy, the addition of Glue5-Azo-SA resulted in deceleration of the hydrolytic reaction (Figure 5c, red), indicating that the SA moiety of Glue5-Azo-SA, even in the cell lysates, selectively binds to the active site of CA as the target enzyme. Of further interest, the hydrolytic reaction was photochemically modulated (Figure 5c, red) in response to the isomerization of the Azo unit of Glue5-Azo-SA in a manner analogous to that in the absence of cell lysates (Figure 5b, red).

Figure 6. Normalized hydrolysis rates of PNPA (1.2 mM) at 37 °C in HEPES buffer (50 mM) containing CA (1 μM) and Glue5-Azo-SA (2.5 μM) at pH 7.2 (red)28 and 5.2 (blue) under alternate irradiations with UV (365 nm, 20 s) and visible light (>400 nm, 5 s). The hydrolysis rates were evaluated as the increment rates of the absorption at 405 nm, which were normalized to that of the initial reaction rate of untreated CA.

Azo-SA conjugate likewise enhanced and attenuated its hydrolytic activity repeatedly in response to the photochemical isomerization of the Azo unit. Such an explicit photochemical response was observable only when the Azo-SA unit (2.5 μM) was attached to Glue5 (Figure 5b, blue). Accordingly, Glue4Azo (2.5 μM) without SA did not suppress the CA-mediated hydrolysis of PNPA (Figure 5b, green), whereas sulfanilamide (2.5 μM; Figure 1e),29 as a noncovalent alternative to the SA moiety, suppressed the reaction considerably (Figure 5b, orange).30 Under acidic conditions at pH 5.2, for example, the negative charges on CA (pI = 5.9)31 are substantially neutralized, so that the Glue5 part of Glue5-Azo-SA would not adhere onto the CA surface. We found that, even at pH 5.2, Glue5-Azo-SA suppressed the hydrolysis of PNPA as a possible consequence of the binding of SA to the active site of CA. However, the PNPA hydrolysis was not modulated (Figure 6, blue; Figure S18)20 in response to the photoisomerization of Glue5-Azo-SA (Figure S10),20 again indicating that the adhesion of the Glue5 part is essential for Glue5-Azo-SA to photomodulate the hydrolytic activity of CA (Figure 2d). As described earlier, the SA moiety of Glue5-Azo-SA docks at the active site of CA (Kassoc = 9.3 ± 3.5 × 107 M−1; Table 1)

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CONCLUSIONS In conclusion, we demonstrated that adhesive azobenzene photoswitch Glue5-Azo-SA, appended at its one terminus with an inhibitor (SA) for CA, serves as a noncovalent photochemical modulator for the enzymatic activity of CA. Glue5Azo-SA can operate even under physiological conditions such as in cell lysates. This achievement, together with the estimated binding affinities, allows us to conclude that the basic design strategy of adhesive photoswitch Gluen-Azo-RInhibit (Figure 2) may operate for other enzymes if appropriate inhibitory motifs (RInhibit) for target enzymes are selected. Considering also the excellent cell-membrane permeabilities of molecular glues,14c selective photochemical modulation of intracellular enzymes with Gluen-Azo-RInhibit is an interesting subject worthy of further investigation for the progress of supramolecular photopharmacology.1f

Table 1. Association (Kassoc) and Dissociation (Kd) Constants of Azo-SA and Glue5-Azo-SA toward CAa

Azo-SA Kassoc (M−1) Kd Glue5-Azo-SA Kassoc (M−1) Kd a

without PNPA (− UV)

with PNPA (− UV)

with PNPA (+ UV)

6.8 ± 3.0 × 107 17 ± 7 nM

4.0 ± 2.1 × 105 2.9 ± 1.4 μM

3.1 ± 0.7 × 105 3.3 ± 0.7 μM

9.3 ± 3.5 × 107 12 ± 4 nM

8.9 ± 0.5 × 105 1.1 ± 0.1 μM

1.7 ± 0.1 × 105 6.0 ± 0.4 μM

The Kassoc and Kd values represent the means ± SD.

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and then possibly guides the connecting Glue5 part to an oxyanion-rich area in proximity to the active site (Kassoc = 2.6 ± 1.4 × 105 M−1; Figure 4b), thereby ensuring the CA/Glue5Azo-SA conjugate (Figure 2a). Considering the Kassoc values corresponding to these events, one may wonder if the Glue5 part is preferentially detached from CA upon photoisomeriza-

ASSOCIATED CONTENT

S Supporting Information *

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/jacs.7b05151. Synthesis of Azo-SA, Gluen-Azo-SA, Gluen-Azo, and Gluen-Cy5.5; 1H NMR, 13C NMR, and MALDI-TOF 10076


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(b) Tochitsky, I.; Banghart, M. R.; Mourot, A.; Yao, J. Z.; Gaub, B.; Kramer, R. H.; Trauner, D. Nat. Chem. 2012, 4, 105−111. (c) Barber, D. M.; Schönberger, M.; Burgstaller, J.; Levitz, J.; Weaver, C. D.; Isacoff, E. Y.; Trauner, D. Chem. Sci. 2016, 7, 2347−2352. (7) Tsai, Y.-H.; Essig, S.; James, J. R.; Lang, K.; Chin, J. W. Nat. Chem. 2015, 7, 554−561. (8) Examples of noncovalent photoswitches for neurotransmitter receptors, see: (a) Volgraf, M.; Gorostiza, P.; Szobota, S.; Helix, M. R.; Isacoff, E. Y.; Trauner, D. J. Am. Chem. Soc. 2007, 129, 260−261. (b) Stawski, P.; Sumser, M.; Trauner, D. Angew. Chem., Int. Ed. 2012, 51, 5748−5751. (c) Stein, M.; Middendorp, S. J.; Carta, V.; Pejo, E.; Raines, D. E.; Forman, S. A.; Sigel, E.; Trauner, D. Angew. Chem., Int. Ed. 2012, 51, 10500−10504. (d) Stein, M.; Breit, A.; Fehrentz, T.; Gudermann, T.; Trauner, D. Angew. Chem., Int. Ed. 2013, 52, 9845− 9848. (e) Schönberger, M.; Trauner, D. Angew. Chem., Int. Ed. 2014, 53, 3264−3267. (f) Laprell, L.; Repak, E.; Franckevicius, V.; Hartrampf, F.; Terhag, J.; Hollmann, M.; Sumser, M.; Rebola, N.; DiGregorio, D. A.; Trauner, D. Nat. Commun. 2015, 6, 8076−8086. (9) Examples of noncovalent photoswitches for ion channels, see: (a) Schoenberger, M.; Damijonaitis, A.; Zhang, Z.; Nagel, D.; Trauner, D. ACS Chem. Neurosci. 2014, 5, 514−518. (b) Schönberger, M.; Althaus, M.; Fronius, M.; Clauss, W.; Trauner, D. Nat. Chem. 2014, 6, 712−719. (c) Broichhagen, J.; Schönberger, M.; Cork, S. C.; Frank, J. A.; Marchetti, P.; Bugliani, M.; Shapiro, A. M. J.; Trapp, S.; Rutter, G. A.; Hodson, D. J.; Trauner, D. Nat. Commun. 2014, 5, 5116−5126. (d) Barber, D. M.; Schönberger, M.; Burgstaller, J.; Levitz, J.; Weaver, C. D.; Isacoff, E. Y.; Baier, H.; Trauner, D. Chem. Sci. 2016, 7, 2347− 2352. (10) Borowiak, M.; Nahaboo, W.; Reynders, M.; Nekolla, K.; Jalinot, P.; Hasserodt, J.; Rehberg, M.; Delattre, M.; Zahler, S.; Vollmar, A.; Trauner, D.; Thorn-Seshold, O. Cell 2015, 162, 403−411. (11) (a) Velema, W. A.; van der Berg, J. P.; Hansen, M. J.; Szymanski, W.; Driessen, A. J. M.; Feringa, B. L. Nat. Chem. 2013, 5, 924−928. (b) Broichhagen, J.; Jurastow, I.; Iwan, K.; Kummer, W.; Trauner, D. Angew. Chem., Int. Ed. 2014, 53, 7657−7660. (c) Hansen, M. J.; Velema, W. A.; de Bruin, G.; Overkleeft, H. S.; Szymanski, W.; Feringa, B. L. ChemBioChem 2014, 15, 2053−2057. (d) Szymanski, W.; Ourailidou, M. E.; Velema, W. A.; Dekker, F. J.; Feringa, B. L. Chem. Eur. J. 2015, 21, 16517−16524. (e) Reis, S. A.; Ghosh, B.; Hendricks, J. A.; Szantai-Kis, D. M.; Törk, L.; Ross, K. N.; Lamb, J.; Read-Button, W.; Zheng, B.; Wang, H.; Salthouse, C.; Haggarty, S. J.; Mazitschek, R. Nat. Chem. Biol. 2016, 12, 317−323. (12) Supuran, C. T.; Scozzafava, A.; Casini, A. Med. Res. Rev. 2003, 23, 146−189. (13) Examples of dendritic molecular glues, see: (a) Okuro, K.; Kinbara, K.; Tsumoto, K.; Ishii, N.; Aida, T. J. Am. Chem. Soc. 2009, 131, 1626−1627. (b) Wang, Q.; Mynar, J. L.; Yoshida, M.; Lee, E.; Lee, M.; Okuro, K.; Kinbara, K.; Aida, T. Nature 2010, 463, 339−343. (c) Okuro, K.; Kinbara, K.; Takeda, K.; Inoue, Y.; Ishijima, A.; Aida, T. Angew. Chem., Int. Ed. 2010, 49, 3030−3033. (d) Suzuki, Y.; Okuro, K.; Takeuchi, T.; Aida, T. J. Am. Chem. Soc. 2012, 134, 15273−15276. (e) Uchida, N.; Okuro, K.; Niitani, Y.; Ling, X.; Ariga, T.; Tomishige, M.; Aida, T. J. Am. Chem. Soc. 2013, 135, 4684−4687. (f) Garzoni, M.; Okuro, K.; Ishii, N.; Aida, T.; Pavan, G. M. ACS Nano 2014, 8, 904− 914. (14) Examples of linear molecular glues, see: (a) Tamesue, S.; Ohtani, M.; Yamada, K.; Ishida, Y.; Spruell, J. M.; Lynd, N. A.; Hawker, C. J.; Aida, T. J. Am. Chem. Soc. 2013, 135, 15650−15655. (b) Mogaki, R.; Okuro, K.; Aida, T. Chem. Sci. 2015, 6, 2802−2805. (c) Hashim, P. K.; Okuro, K.; Sasaki, S.; Hoashi, Y.; Aida, T. J. Am. Chem. Soc. 2015, 137, 15608−15611. (d) Hatano, J.; Okuro, K.; Aida, T. Angew. Chem., Int. Ed. 2016, 55, 193−198. (e) Okuro, K.; Sasaki, M.; Aida, T. J. Am. Chem. Soc. 2016, 138, 5527−5530. (15) Examples of guanidinium-rich polymers, see: (a) Sakai, N.; Matile, S. J. Am. Chem. Soc. 2003, 125, 14348−14356. (b) Hennig, A.; Gabriel, G. J.; Tew, G. N.; Matile, S. J. Am. Chem. Soc. 2008, 130, 10338−10344. (c) Shukla, D.; Schneider, C. P.; Trout, B. L. J. Am. Chem. Soc. 2011, 133, 18713−18718. (d) Geihe, E. I.; Cooley, C. B.; Simon, J. R.; Kiesewetter, M. K.; Edward, J. A.; Hickerson, R. P.;

mass spectral data; electronic absorption spectra; and related experimental procedures (PDF)

AUTHOR INFORMATION

Corresponding Authors

*[email protected] *[email protected] ORCID

Kou Okuro: 0000-0003-0445-6358 Takuzo Aida: 0000-0002-0002-8017 Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS This work was partially supported by a Grant-in-Aid for Young Scientists (B) (26810046) to K.O. R.M. thanks the Research Fellowships of Japan Society for the Promotion of Science (JSPS) for Young Scientists and the Program for Leading Graduate Schools (GPLLI).

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REFERENCES

(1) For reviews of spatiotemporal modulation of biological processes, see: (a) Mayer, G.; Heckel, A. Angew. Chem., Int. Ed. 2006, 45, 4900− 4921. (b) Young, D. D.; Deiters, A. Org. Biomol. Chem. 2007, 5, 999− 1005. (c) Lee, H.-M.; Larson, D. R.; Lawrence, D. S. ACS Chem. Biol. 2009, 4, 409−427. (d) Deiters, A. ChemBioChem 2010, 11, 47−53. (e) Fehrentz, T.; Schönberger, M.; Trauner, D. Angew. Chem., Int. Ed. 2011, 50, 12156−12182. (f) Lerch, M. M.; Hansen, M. J.; van Dam, G. M.; Szymanski, W.; Feringa, B. L. Angew. Chem., Int. Ed. 2016, 55, 10978−10999. (2) For reviews of photoswitches, see: (a) Gorostiza, P.; Isacoff, E. Mol. BioSyst. 2007, 3, 686−704. (b) Gorostiza, P.; Isacoff, E. Y. Science 2008, 322, 395−399. (c) Riggsbee, C. W.; Deiters, A. Trends Biotechnol. 2010, 28, 468−475. (d) Beharry, A. A.; Woolley, G. A. Chem. Soc. Rev. 2011, 40, 4422−4437. (e) Szymański, W.; Beierle, J. M.; Kistemaker, H. A. V.; Velema, W. A.; Feringa, B. L. Chem. Rev. 2013, 113, 6114−6178. (f) Broichhagen, J.; Trauner, D. Curr. Opin. Chem. Biol. 2014, 21, 121−127. (3) Bartels, E.; Wassermann, N. H.; Erlanger, B. F. Proc. Natl. Acad. Sci. U. S. A. 1971, 68, 1820−1823. (4) Examples of covalently tethered photoswitches on potassium ion channels, see: (a) Banghart, M.; Borges, K.; Isacoff, E.; Trauner, D.; Kramer, R. H. Nat. Neurosci. 2004, 7, 1381−1386. (b) Chambers, J. J.; Banghart, M. R.; Trauner, D.; Kramer, R. H. J. Neurophysiol. 2006, 96, 2792−2796. (c) Fortin, D. L.; Banghart, M. R.; Dunn, T. W.; Borges, K.; Wagenaar, D. A.; Gaudry, Q.; Karakossian, M. H.; Otis, T. S.; Kristan, W. B.; Trauner, D.; Kramer, R. H. Nat. Methods 2008, 5, 331− 338. (5) Examples of covalently tethered photoswitches on iGluR, see: (a) Volgraf, M.; Gorostiza, P.; Numano, R.; Kramer, R. H.; Isacoff, E. Y.; Trauner, D. Nat. Chem. Biol. 2006, 2, 47−52. (b) Szobota, S.; Gorostiza, P.; Bene, F. D.; Wyart, C.; Fortin, D. L.; Kolstad, K. D.; Tulyathan, O.; Volgraf, M.; Numano, R.; Aaron, H. L.; Scott, E. K.; Kramer, R. H.; Flannery, J.; Baier, H.; Trauner, D.; Isacoff, E. Y. Neuron 2007, 54, 535−545. (c) Numano, R.; Szobota, S.; Lau, A. Y.; Gorostiza, P.; Volgraf, M.; Roux, B.; Trauner, D.; Isacoff, E. Y. Proc. Natl. Acad. Sci. U. S. A. 2009, 106, 6814−6819. (d) Wyart, C.; Bene, F. D.; Warp, E.; Scott, E. K.; Trauner, D.; Baier, H.; Isacoff, E. Y. Nature 2009, 461, 407−410. (e) Caporale, N.; Kolstad, K. D.; Lee, T.; Tochitsky, I.; Dalkara, D.; Trauner, D.; Kramer, R.; Dan, Y.; Isacoff, E. Y.; Flannery, J. G. Mol. Ther. 2011, 19, 1212−1219. (f) Polosukhina, A.; Litt, J.; Tochitsky, I.; Nemargut, J.; Sychev, Y.; De Kouchkovsky, I.; Huang, T.; Borges, K.; Trauner, D.; Van Gelder, R. N.; Kramer, R. H. Neuron 2012, 75, 271−282. (6) Examples of other covalently tethered photoswitches, see: (a) Harvey, J. H.; Trauner, D. ChemBioChem 2008, 9, 191−193. 10077


Article

Journal of the American Chemical Society Kaspar, R. L.; Hedrick, J. L.; Waymouth, R. M.; Wender, P. A. Proc. Natl. Acad. Sci. U. S. A. 2012, 109, 13171−13176. (e) Yonamine, Y.; Yoshimatsu, K.; Lee, S.-H.; Hoshino, Y.; Okahata, Y.; Shea, K. J. ACS Appl. Mater. Interfaces 2013, 5, 374−379. (f) Bang, E.-K.; Gasparini, G.; Molinard, G.; Roux, A.; Sakai, N.; Matile, S. J. Am. Chem. Soc. 2013, 135, 2088−2091. (g) Gasparini, G.; Bang, E.-K.; Molinard, G.; Tulumello, D. V.; Ward, S.; Kelley, S. O.; Roux, A.; Sakai, N.; Matile, S. J. Am. Chem. Soc. 2014, 136, 6069−6074. (h) Gasparini, G.; Matile, S. Chem. Commun. 2015, 51, 17160−17162. (i) Chuard, N.; Gasparini, G.; Roux, A.; Sakai, N.; Matile, S. Org. Biomol. Chem. 2015, 13, 64−67. (j) McKinlay, C. J.; Waymouth, R. M.; Wender, P. A. J. Am. Chem. Soc. 2016, 138, 3510−3517. (k) Morelli, P.; Martin-Benlloch, X.; Tessier, R.; Waser, J.; Sakai, N.; Matile, S. Polym. Chem. 2016, 7, 3465−3470. (16) Modified image of 1V9E: Saito, R.; Sato, T.; Ikai, A.; Tanaka, N. Acta Crystallogr., Sect. D: Biol. Crystallogr. 2004, 60, 792−795. Created with PyMOL 1.3. (17) (a) Banerjee, A. L.; Swanson, M.; Roy, B. C.; Jia, X.; Haldar, M. K.; Mallik, S.; Srivastava, D. K. J. Am. Chem. Soc. 2004, 126, 10875− 10883. (b) Roy, B. C.; Banerjee, A. L.; Swanson, M.; Jia, X. G.; Haldar, M. K.; Mallik, S.; Srivastava, D. K. J. Am. Chem. Soc. 2004, 126, 13206−13207. (c) Jude, K. M.; Banerjee, A. L.; Haldar, M. K.; Manokaran, S.; Roy, B.; Mallik, S.; Srivastava, D. K.; Christianson, D. W. J. Am. Chem. Soc. 2006, 128, 3011−3018. (18) For reviews of click reaction, see: (a) Kolb, H. C.; Finn, M. G.; Sharpless, K. B. Angew. Chem., Int. Ed. 2001, 40, 2004−2021. (b) Díaz, D. D.; Punna, S.; Holzer, P.; McPherson, A. K.; Sharpless, K. B.; Fokin, V. V.; Finn, M. G. J. Polym. Sci., Part A: Polym. Chem. 2004, 42, 4392− 4403. (19) Examples of synthesis of functional polymers by click reaction, see: (a) Gong, T.; Adzima, B. J.; Baker, N. H.; Bowman, C. N. Adv. Mater. 2013, 25, 2024−2028. (b) Accurso, A. A.; Delaney, M.; O’Brien, J.; Kim, H.; Iovine, P. M.; Díaz, D. D.; Finn, M. G. Chem. Eur. J. 2014, 20, 10710−10719. (c) Sandmann, B.; Happ, B.; Hager, M. D.; Vitz, J.; Rettler, E.; Burtscher, P.; Moszner, N.; Schubert, U. S. J. Polym. Sci., Part A: Polym. Chem. 2014, 52, 239−247. (20) See Supporting Information. (21) Schobel, U.; Egelhaaf, H.-J.; Brecht, A.; Oelkrug, D.; Gauglitz, G. Bioconjugate Chem. 1999, 10, 1107−1114. (22) Yi, L.; Morgan, J. T.; Ragsdale, S. W. J. Biol. Chem. 2010, 285, 20117−20127. (23) Chen, R. F.; Kernohan, J. C. J. Biol. Chem. 1967, 242, 5813− 5823. (24) (a) Nair, S. K.; Krebs, J. F.; Christianson, D. W.; Fierke, C. A. Biochemistry 1995, 34, 3981−3989. (b) Wang, S. C.; Zamble, D. B. Biochem. Mol. Biol. Educ. 2006, 34, 364−368. (25) Vomasta, D.; Högner, C.; Branda, N. R.; König, B. Angew. Chem., Int. Ed. 2008, 47, 7644−7647. (26) (a) Geers, C.; Gros, G. Physiol. Rev. 2008, 7, 681−715. (b) Supuran, C. T. Nat. Rev. Drug Discovery 2008, 7, 168−181. (c) Casey, J. R.; Grinstein, S.; Orlowski, J. Nat. Rev. Mol. Cell Biol. 2010, 11, 50−61. (27) Pocker, Y.; Stone, J. T. Biochemistry 1967, 6, 668−678. (28) The gradual decrease in hydrolysis rate is due to the consumption of PNPA. (29) Mann, T.; Keilin, D. Nature 1940, 146, 164−165. (30) Nevertheless, no photochemical response resulted, thereby excluding a possible contribution of the photothermal effect of the excited Azo unit to the modulation of the hydrolytic reaction. (31) Barbet, A. F.; Blentlinger, R.; Yi, J.; Lundgren, A. M.; Blouin, E. F.; Kocan, K. M. Infect. Immun. 1999, 67, 102−107.

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