Geometry-Induced Oligomerization of Fluorine-Substituted Phenylazothiazole Photoswitches.

Photoswitches are molecules that can absorb light of specific wavelengths and undergo a reversible transformation between their trans and cis isomeric forms. In phenylazo photoswitches, it is common for the less stable cis (Z) isomer to convert back to the more stable trans (E) isomer either through photochemical or thermal means. In this research, we designed new derivatives of phenylazothiazole (PAT) photoswitches, PAT-Fn, which feature fluorine substituents on their phenyl component. These derivatives can reversibly isomerize under visible light exposure with the enrichment of E and Z isomers at photostationary state (PSS). Surprisingly, we observed an unconventional phenomenon when these PAT-Fn (n≧2) photoswitches were in their cis isomeric state in the absence of light. Instead of the anticipated transformation from cis to trans isomer, these compounds converted to an oligomeric compound. Our detailed experimental investigation and theoretical calculations, indicated the crucial role of fluorine substituents and the distinctive geometric arrangement of the cis isomer in driving the unexpected oligomerization process originating from the cis isomeric state.

Chaperonin GroEL hydrolyses ortho-nitrophenyl ß-galactoside.

We serendipitously found that chaperonin GroEL can hydrolyze ortho-nitrophenyl ß-galactoside (ONPG), a well-known substrate of the enzyme ß-galactosidase. The ONPG hydrolysis by GroEL follows typical enzyme kinetics. Our experiments and molecular docking studies suggest ONPG binding at the ATP binding site of GroEL.

Phenylazothiazoles as Visible-Light Photoswitches.

A novel class of photoswitches based on a phenylazothiazole scaffold that undergoes reversible isomerization under visible-light irradiation is reported. The photoswitch, which comprises a thiazole heteroaryl segment directly connected to a phenyl azo chromophore, has very different spectral characteristics, such as a redshifted absorption maximum wavelength and well-separated absorption bands of the trans and cis isomers, than conventional azobenzene and other heteroaryl azo compounds. Substituents at the ortho and para positions of the phenyl ring of the photoswitch resulted in a further shift to longer wavelengths up to 525 nm at the absorption maximum with a small thermal stability compensation. These photoswitches showed excellent photostationary distributions of the trans and cis isomers, thermal half-lives of up to 7.2 h, and excellent reductant stability. The X-ray crystal structure analysis revealed that the trans isomers exhibited a planar geometry and the cis isomers exhibited a T-shaped orthogonal geometry. Detailed ab initio calculations further demonstrated the plausible electronic transitions and isomerization energy barriers, which were consistent with the experimental observations. The fundamental design principles elucidated in this study will aid in the development of a wide variety of visible-light photoswitches for photopharmacological applications.

Tetraploidy-linked sensitization to CENP-E inhibition in human cells.

Tetraploidy is a hallmark of cancer cells, and tetraploidy-selective cell growth suppression is a potential strategy for targeted cancer therapy. However, how tetraploid cells differ from normal diploids in their sensitivity to anti-proliferative treatments remains largely unknown. In this study, we found that tetraploid cells are significantly more susceptible to inhibitors of a mitotic kinesin (CENP-E) than are diploids. Treatment with a CENP-E inhibitor preferentially diminished the tetraploid cell population in a diploid-tetraploid co-culture at optimum conditions. Live imaging revealed that a tetraploidy-linked increase in unsolvable chromosome misalignment caused substantially longer mitotic delay in tetraploids than in diploids upon moderate CENP-E inhibition. This time gap of mitotic arrest resulted in cohesion fatigue and subsequent cell death, specifically in tetraploids, leading to tetraploidy-selective cell growth suppression. In contrast, the microtubule-stabilizing compound paclitaxel caused tetraploidy-selective suppression through the aggravation of spindle multipolarization. We also found that treatment with a CENP-E inhibitor had superior generality to paclitaxel in its tetraploidy selectivity across a broader spectrum of cell lines. Our results highlight the unique properties of CENP-E inhibitors in tetraploidy-selective suppression and their potential use in the development of tetraploidy-targeting interventions in cancer.

Excited State Charge-Transfer Complexes Enable Fluorescence Color Changes in a Supramolecular Cyclophane Mechanophore.

Mechanochromic mechanophores are reporter molecules that indicate mechanical events through changes of their photophysical properties. Supramolecular mechanophores in which the activation is based on the rearrangement of luminophores and/or quenchers without any covalent bond scission, remain less well investigated. Here, we report a cyclophane-based supramolecular mechanophore that contains a 1,6-bis(phenylethynyl)pyrene luminophore and a pyromellitic diimide quencher. In solution, the blue monomer emission of the luminophore is largely quenched and a faint reddish-orange emission originating from a charge-transfer (CT) complex is observed. A polyurethane elastomer containing the mechanophore displays orange emission in the absence of force, which is dominated by the CT-emission. Mechanical deformation causes a decrease of the CT-emission and an increase of blue monomer emission, due to the spatial separation between the luminophore and quencher. The ratio of the two emission intensities correlates with the applied stress.

Dynamic Control of Microbial Movement by Photoswitchable ATP Antagonists.

Adenosine triphosphate (ATP) is the energy source for various biochemical processes and biomolecular motors in living things. Development of ATP antagonists and their stimuli-controlled actions offer a novel approach to regulate biological processes. Herein, we developed azobenzene-based photoswitchable ATP antagonists for controlling the activity of motor proteins; cytoplasmic and axonemal dyneins. The new ATP antagonists showed reversible photoswitching of cytoplasmic dynein activity in an in vitro dynein-microtubule system due to the trans and cis photoisomerization of their azobenzene segment. Importantly, our ATP antagonists reversibly regulated the axonemal dynein motor activity for the force generation in a demembranated model of Chlamydomonas reinhardtii. We found that the trans and cis isomers of ATP antagonists significantly differ in their affinity to the ATP binding site.

A visible light-controllable Rho kinase inhibitor based on a photochromic phenylazothiazole.

Rho-associated coiled-coil-containing protein kinase (ROCK) is a serine-threonine kinase whose inhibitors are useful for the regulation of the actomyosin system. Here, we developed a photoswitchable ROCK inhibitor based on a phenylazothiazole scaffold. The reversible trans-cis isomerization by visible light stimuli enabled us to manipulate ROCK activities in vitro and in cells.

Synthesis and Properties of Aromatic-Terminated Diacetylene Organogelators and Their Application to Photopatterning of Polydiacetylenes.

A series of simply structured diacetylene-diamide-based gelators (DAGs) with aromatic terminals were synthesized, and their gelation and subsequent photopolymerization abilities were analyzed. DAGs with an adequate spacer length (n) and tolyl terminals (DA-Tn) interacted with aromatic solvents, such as benzene and xylenes, at elevated temperatures. During the subsequent cooling process, the DAGs interacted with each other through CH-π interactions at their terminal positions. They also formed one-dimensional hydrogen bonding arrays through secondary amides, leading to stable organogels. These gels polymerized into π-conjugated polydiacetylenes (PDAs) under ultraviolet irradiation. In the p-xylene gels of DA-Tn, the spacer length exerted characteristic odd-even effects on the photopolymerization rates over a certain range (n = 3-6), which can be explained by periodic changes in the uniformity of the molecular packing modes. When the gelling solvent was changed to cyclohexane, the gelation and photopolymerization abilities were greatly improved because the DA-Tn gel networks became highly crystallized and transparent to ultraviolet light (254 nm). The ultimate conversion to PDA from DA-T8/cyclohexane gels was 45.2 wt %. Applying photolithographic techniques to the DAG with excellent photopolymerizability in the film state, we successfully fabricated microscale photopatterns of PDA. We also established a convenient removal process (development process) of DA monomers in unexposed areas. The resulting PDA patterns were quite stable to ambient light stimuli.

Rational design and development of a lit-active photoswitchable inhibitor targeting CENP-E.

In the emerging field of photopharmacology, synthetic photoswitches based on reversible photochemical reactions are fused to bioactive molecules. Azobenzene derivatives, which can undergo trans-cis photoisomerization, are typical photoswitches. Most azobenzene-based photochemical tools are active in the thermodynamically stable trans, but not cis, form. cis-Active photochemical tools would be ideal because they can be "initially inactive and active after light illumination" in a reversible mode only by light illumination. However, only a few rational strategies for constructing such "lit-active" photopharmacological tools has been developed. Herein, we report a rationally designed lit-active photoswitchable inhibitor targeting centromere-associated protein E (CENP-E). Using the lit-active inhibitor, we were able to photoregulate CENP-E-dependent mitotic chromosome location in cells. This study provides a framework to facilitate further progress in the development of photopharmacological tools.

Rotaxane-Based Dual Function Mechanophores Exhibiting Reversible and Irreversible Responses.

Mechanochromic mechanophores permit the design of polymers that indicate mechanical events through optical signals. Here we report rotaxane-based supramolecular mechanophores that display both reversible and irreversible fluorescence changes. These responses are triggered by different forces and are achieved by exploiting the molecular shuttling function and force-induced dethreading of rotaxanes. The new rotaxane mechanophores are composed of a ring featuring a luminophore, which is threaded onto an axle with a matching quencher and two stoppers. In the stress-free state, the luminophore is preferentially located in the proximity of the quencher, and the emission is quenched. The luminophore slides away from the quencher when a force is applied and the fluorescence is switched on. This effect is reversible, unless the force is so high that the luminophore-carrying ring slips past the stopper and dethreading occurs. We show that the combination of judiciously selected ring and stopper moieties is crucial to attain interlocked structures that display such a dual response. PU elastomers that contain such doubly responsive rotaxanes exhibit reversible fluorescence changes over multiple loading-unloading cycles due to the shuttling function, whereas permanent changes are observed upon repeated deformations to high strains due to breakage of the mechanical bond upon dethreading of the ring from the axle. This response allows one, at least conceptually, to monitor the actual deformation of polymer materials and examine mechanical damage that was inflicted in the past on the basis of an optical signal.

Mechanically Responsive Luminescent Polymers Based on Supramolecular Cyclophane Mechanophores.

A new approach to cyclophane-based supramolecular mechanophores is presented. We report a mechanically responsive cyclic motif that contains two fluorescent 1,6-bis(phenylethynyl)pyrene moieties that are capable of forming intramolecular excimers. The emission spectra of dilute solutions of this cyclophane and a polyurethane elastomer into which a small amount of the mechanophore (0.08 wt %) had been covalently integrated are dominated by excimer emission. Films of the cyclophane-containing polyurethane also display a considerable portion of excimer emission, but upon deformation, the fluorescence becomes monomer-dominated and a perceptible change from cyan to blue is observed. The response is instant, reversible, and consistent with a mechanically induced change of the molecular conformation of the mechanophore so that the excimer-promoting interactions between the luminophores are suppressed. In-depth investigations show a correlation between the applied strain and the emission color, which can conveniently be expressed by the ratio of monomer to excimer emission intensity. The current study suggests that cyclophanes can be utilized to develop various supramolecular mechanophores that detect and visualize weak forces occurring in polymeric materials or generated by living tissues.

A Series of Bisamide-Substituted Diacetylenes Exhibiting a Terminal Alkyl Odd/Even Parity Effect on Mechanoactivated Photopolymerization.

Diacetylene derivatives exhibit solid-state polymerization to polydiacetylene initiated by UV light or γ-ray irradiation. The activation of the photopolymerization relies on the monomer diynes arrangement. Recently, it has been demonstrated that the first mechanoresponsive bisamide substituted diacetylenes (DAs) show dramatic switching from light-inert to light-reactive states at a given pressure. The origin of this unique phenomenon was apparently related to the pressure-sensitive crystalline transition in DAs, but the molecular mechanism remains elusive. To obtain more insight, herein a series of DAs with varying terminal alkyl spacer length is presented, and their molecular structural effect on the intermolecular hydrogen bonding and steric repulsion is examined. In pristine states, even-parity DAs were inactive upon UV irradiation (λ=254 nm) unless external pressure was applied. By contrast, odd-parity DAs were easily polymerized upon UV irradiation without pressure application. However, the pressure-induced crystalline phase transition exhibiting photopolymerization was valid for all DAs regardless of their alkyl spacer length. A systematic investigation revealed that the terminal alkyl spacer length, especially its odd/even parity plays a key role in determining the intrinsic intermolecular hydrogen-bonding nature of DA crystals and the resultant molecular packing. In addition, the relevant thermochromic behavior was also observed from photopolymerized polydiacetylenes.

Glycomacrocycle-Based Azobenzene Derivatives as Chiral Dopants for Photoresponsive Cholesteric Liquid Crystals.

We demonstrate photoresponsive cholesteric liquid crystals (CLCs) doped with glycomacrocyclic azobenzene derivatives, which exhibit large conformational changes, providing dynamic control of helical superstructures in response to a light stimulus. An unprecedented shortening of the helical pitch length and the empowerment of helical twisting power up to 500% are observed upon trans (E) to cis (Z) photoisomerization. Light-driven dynamic helix twisting and untwisting behavior affords the first example of glycomacrocyclic azobenzene-based CLCs, which can drive the mechanical movement of micro-objects. Two modes of rotation-two-directional or one-directional rotational motion (crankshaft mode)-are realized. In particular, the latter mode based on the reversible cholesteric texture transition between homogeneous stripes and focal conics leads to the accumulation of the rotation angles achieving the amplified mechanical movements.

Electrofluorochromic Device Based on a Redox-Active Europium(III) Complex.

Electrofluorochromism owing to redox reactions on the center europium (Eu) ion in ionic liquids is examined for the helicate complexes (abbreviated as EuL) with a hexadentate pyridine derivative. Typical electrofluorochromism requires extra electroactive units complementing intra- or intermolecular energy transfer to quench fluorophores. Herein, an unprecedentedly simplified electrofluorochromic system overcoming such issues is demonstrated by utilizing reversible electrochemistry of EuL between Eu3+ and Eu2+, which accompanies large emission transition. A three-electrode electrochemical switching device is facilely prepared with an ionic liquid [BMIM][PF6] and EuL mixture. Benefiting from the stable helical coordinated structure of the ligand in [BMIM][PF6], highly enhanced red fluorescence of EuL with small quantity (≤1 wt %) is utilized. Rapid response and large contrast of luminescence are achieved: the emission is drastically quenched at the reduced state (Eu2+) and it is successfully restored by subsequent oxidation (Eu3+). The reversible fluctuation of excitation and emission spectra of an electrofluorochromic device is achieved in the potential window within ±2 V. The device affords excellent optoelectric properties in terms of well-controlled luminescence switching depending on the applied potentials and its durability. This work paves an efficient and smart way toward Eu luminescence control in optoelectronic devices.

Photoisomerization of azobenzene units drives the photochemical reaction cycles of proteorhodopsin and bacteriorhodopsin analogues.

In this study we substituted the retinal units in proteorhodopsin (PR) and bacteriorhodopsin (BR) with azo chromophores to investigate the mechanism of photoinduced proton pumping in rhodopsins and potentially develop new artificial molecular pumps. We used an indium tin oxide electrode to investigate the photoinduced proton transfer of the azo analogues of PR and BR. We also employed flash photolysis to determine the characteristic photocycles, comprising multiple transient intermediates, of the azo chromophore-bound PR and BR. Moreover, our studies of the photoinduced proton pumping functions of azo-proteoopsin and azo-bacterioopsin complexes revealed that they did not pump protons upon illumination, even though they underwent photoinduced proton transfer and the characteristic photocycle. Mutational analysis suggested that the proton pumping malfunction of the azo analogues of PR and BR resulted from the absence of proton transfer reactions through cytoplasmic channels, even though these reactions were evoked in extracellular channels. Based on our experimental findings, we propose herein a putative model of the proton transfer reaction mechanism for the azo analogues of PR and BR.

Photoswitchable CENP-E Inhibitor Enabling the Dynamic Control of Chromosome Movement and Mitotic Progression.

Interfering with mitosis is a potential cancer therapy strategy. However, the lack of controllability of antimitotic drugs in cell growth suppression causes severe side effects and limits their clinical utility. Herein, we developed an azobenzene-based photoswitchable inhibitor of CENP-E, a mitotic kinesin required for chromosome transportation. The new inhibitor enabled reversible photoswitching of CENP-E activity with ∼10-fold change in IC50 between cis and trans photoisomerization states both in vitro and in living cells. It also enabled repeatable photoswitching of CENP-E-dependent chromosome congression and hence mitotic progression with UV/vis light illumination cycles. Using this technique, we could specify the exact process of mitotic progression in which CENP-E plays an indispensable role. Our data demonstrate the power of a photochemical approach for highly controllable mitotic interference as well as for discovery of precise molecular functions in dynamic cellular processes.

Mechanochromic Luminescence from Crystals Consisting of Intermolecular Hydrogen-Bonded Sheets.

Introduction of functional groups that can form intermolecular hydrogen bonds into highly-emissive luminophores is a promising way to induce mechanochromic luminescence. Herein, we report that a 9,10-bis(phenylethynyl)anthracene derivative featuring two amide groups forms green-emissive crystals based on two-dimensional hydrogen-bonded molecular sheets. Mechanical grinding changed the emission from green to yellow, owing to a transition from a crystalline to an amorphous phase. Infrared spectroscopy revealed that mechanical stimuli disrupted the linear hydrogen-bonding formation. A thermal treatment recovered the original green photoluminescence.

Mechanoresponsive Behavior of a Polymer-Embedded Red-Light Emitting Rotaxane Mechanophore.

A red light-emitting photoluminescent supramolecular mechanophore based on an interlocked molecular motif is presented. The rotaxane-based mechanophore contains a cyclic compound featuring a π-extended 4,4-difluoro-4-bora-3a,4a-diaza-s-indacene (BODIPY) dye as a red emitter that was threaded onto a dumbbell-shaped molecule containing an electron-poor 1,4,5,8-naphthalenetetracarboxylic diimide quencher at its center. Through two aliphatic hydroxyl groups attached to the dumbbell and the cycle, the mechanophore was covalently embedded into the backbone of a thermoplastic polyurethane elastomer. The mechanophore is only weakly photoluminescent in solution, indicating that the BODIPY's emission is efficiently quenched. Solution-cast films of the rotaxane-containing polymer, by contrast, show an appreciable photoluminescence, which suggests that during film formation, some of the emitting cycles are trapped in positions away from the quencher. Interestingly, the emission intensity could be significantly reduced by swelling the films with an organic solvent and the emission increased again upon drying, suggesting that such solvent plasticization causes a reversible rearrangement. In both dry and solvent-swollen films, uniaxial deformation caused a significant, reversible increase of the emission intensity, on account of mechanically induced shuttling of the emitters away from and back to the quenchers. It is shown that the properties of the polymer can be tuned by the solvent, and that such plasticizing extends the small palette of approaches that allow modification of the activation stress of a given material system.

Rotaxane-Based Mechanophores Enable Polymers with Mechanically Switchable White Photoluminescence.

Three mechanoresponsive polyurethane elastomers whose blue, green, and orange photoluminescence can be reversibly turned on by mechanical force were prepared and combined to create a blend that exhibits deformation-induced white photoluminescence. The three polyurethanes contain rotaxane-based supramolecular mechanoluminophores based on π-extended pyrene, anthracene, or 4-(dicyanomethylene)-2-methyl-6-(4-dimethylaminostyryl)-4H-pyran (DCM) luminophores, respectively, and 1,4,5,8-naphthalenetetracarboxylic diimide as an electronically matched quencher. Each polymer shows instantly reversible, strain-dependent switching of its photoluminescence intensity when stretched and relaxed, as deformation leads to a spatial separation of the luminophore and quencher. The present study shows that the photoluminescence color can easily be tailored by variation of the luminophore and also by combining several mechanophores in one material and demonstrates that adaptability is a key advantage of supramolecular approaches to create mechanoresponsive polymers.

Molecular Crankshaft Effect Converting Piston-like Molecular Motion to Continuous Rotation of Macro Objects.

A molecular system inducing an accumulative unidirectional rotation motion of glass flakes with about 100 µm in size is introduced. The molecular system is a chiral nematic liquid crystal containing a chiral azobenzene derivative which shows a reversible E- Z photoisomerization accompanying a large helical twisting power change. A film of the molecular system shows different texture change paths upon UV and visible light irradiations inducing " E to Z" and " Z to E" photoisomerizations, respectively, of the chiral azobenzene dopant. Namely, a polygonal fingerprint texture inducing the rotation of glass flakes on the film surface was maintained during UV irradiation, while a focal conic texture inducing no rotation of glass flakes emerged during visible light irradiation. As a result, cycles of the alternative irradiation of UV and visible lights afforded many rotations toward a single direction of the glass flakes which can be considered as a continuous conversion of light energy to mechanical work. We may compare the effect of this molecular system converting "back and forth" structural change between E and Z isomers of the chiral azobenzene to a continuous rotational motion of glass flakes with the crankshaft effect converting a piston-like motion to a rotational motion seen in engines in the real world.