Takahiro Muraoka

Mechanical twisting of a guest by a photoresponsive host

Molecular analogues of a variety of mechanical devices such as shuttles, brakes, unidirectional rotors and tweezers have been created. But these ‘molecular machines’ have not yet been used to mechanically manipulate a second molecule in a controlled and reversible manner. Here we show that light-induced scissor-like conformational changes of one molecule can give rise to mechanical twisting of a non-covalently bound guest molecule. To realize this coupling of molecular motions, we use a previously designed system: a ferrocene moiety with an azobenzene strap, each end of which is attached to one of the two cyclopentadienyl rings of the ferrocene unit, acts as a pivot so that photoisomerization of the strap rotates the ferrocene rings relative to each other and thereby also changes the relative position of two ‘pedal’ moieties attached to the ferrocene rings. We translate this effect into intermolecular coupling of motion by endowing the pedals with binding sites, which allow the host system to form a stable complex with a bidentate rotor molecule. Using circular dichroism spectroscopy, we show that the photoinduced conformational changes of the host are indeed transmitted and induce mechanical twisting of the rotor molecule. This design concept, which significantly extends the successful coupling of motion beyond the intramolecular level seen in synthetic allosteric receptors, might allow for the remote control of molecular events in larger interlocked molecular systems.

Light‐Triggered Bioactivity in Three Dimensions

Hydrogelators that undergo stimuli-responsive sol–gel transitions have attracted attention as biomaterials because of their potential applications, for example as sophisticated cell culture substrates, drug carriers, microvalves, and microactuators. 2] Among the various stimuli possible, light is unique because it allows both spatial and temporal control of a specific reaction without requiring physical contact. Many photochemical sol–gel transitions have been reported, mostly in polymers but also in low-molecular-weight peptides. 3] The importance of peptide systems is their potential biocompatibility and the opportunity for researchers to molecularly design bioactive functions. The advent of two-dimensional self-assembling systems and patterning technologies has generated some examples of stimulus-driven bioactivity on surfaces. Using a peptide amphiphile (PA) containing the fibronectin epitope Arg-Gly-Asp-Ser (RGDS) for cell adhesion, we demonstrate here light-triggered enhancement of bioactivity in a three-dimensional system. PAs are known as highly versatile molecules that selfassemble into nanostructures such as spherical micelles, fibers, and helices through hydrogen-bond formation and hydrophobic collapse. 5, 8–10] Our laboratory first reported on PAs that form high-aspect-ratio nanofibers and therefore gels at very low concentrations. These specific PAs self-assemble into nanofibers as a result of their b-sheet peptide domains, and their self-assembly can be triggered by charge screening through changes in pH or the addition of salts. Molecular changes in the b-sheet domains can disrupt nanofiber formation. For example, Hartgerink and co-workers reported the formation of spherical micelles and not fibers as a result of the N-methylation on the amide nitrogen closest to the alkyl tail of a PA. Our laboratory also reported recently the formation of quadruple helices in PAs containing a photolabile 2-nitrobenzyl group in the b-sheet domain. These quadruple helices in turn dissociate into single nonhelical fibrils in response to light. In this work, we have synthesized PA molecule 1 containing both the photocleavable 2-nitrobenzyl group as well as the bioactive epitope Arg-Gly-Asp-Ser (RGDS) (Scheme 1). Based on the previous work, photoirradiation

Quadruple Helix Formation of a Photoresponsive Peptide Amphiphile and Its Light-Triggered Dissociation into Single Fibers

Using a peptide amphiphile having a bulky photolabile 2-nitrobenzyl group between the alkyl chain and the peptide segment, we demonstrated quadruple helical fiber formation and its dissociation into single fibrils in response to light. Putting the bulky group close to the core of a fibril is thought to induce a distortion of the alignment of molecules, which can in turn lead to quadruple helices. Photoirradiation to cleave the bulky group transforms the helices into single fibrils.

A Structured Monodisperse PEG for the Effective Suppression of Protein Aggregation

Part of the solution: A PEG with a discrete triangular structure exhibits hydrophilicity/hydrophobicity switching upon increasing temperatures, and suppresses the thermal aggregation of lysozyme to retain nearly 80 % of the enzymatic activity. CD and NMR spectroscopic studies revealed that, with the structured PEG, the higher-order structures of lysozyme persist at high temperature, and the native conformation is recovered after cooling.

Thermally driven polymorphic transition prompting a naked-eye-detectable bending and straightening motion of single crystals.

The amplification of molecular motions so that they can be detected by the naked eye (10(7) -fold amplification from the ångström to the millimeter scale) is a challenging issue in the development of mechanical molecular devices. In this context, the perfectly ordered molecular alignment of the crystalline phase has advantages, as demonstrated by the macroscale mechanical motions of single crystals upon the photochemical transformation of molecules. In the course of our studies on thermoresponsive amphiphiles containing tetra(ethylene glycol) (TEG) moieties, we serendipitously found that thermal conformational changes of TEG units trigger a single-crystal-to-single-crystal polymorphic phase transition. The single crystal of the amphiphile undergoes bending and straightening motion during both heating and cooling processes at the phase-transition temperatures. Thus, the thermally triggered conformational change of PEG units may have the advantage of inducing mechanical motion in bulk materials.

Ion permeation by a folded multiblock amphiphilic oligomer achieved by hierarchical construction of self-assembled nanopores.

A multiblock amphiphilic molecule 1, with a tetrameric alternating sequence of hydrophilic and hydrophobic units, adopts a folded structure in a liposomal membrane like a multipass transmembrane protein, and is able to transport alkali metal cations through the membrane. Hills analysis and conductance measurements, analyzed by the Hille equation, revealed that the tetrameric assembly of 1 forms a 0.53 nm channel allowing for permeation of cations. Since neither 3, bearing flexible hydrophobic units and forming no stacked structures in the membrane, nor 2, a monomeric version of 1, is able to transport cations, the folded conformation of 1 in the membrane is likely essential for realizing its function. Thus, function and hierarchically formed higher-order structures of 1, is strongly correlated with each other like proteins and other biological macromolecules.

Reversible ion transportation switch by a ligand-gated synthetic supramolecular ion channel.

Inspired by the regulation of cellular activities found in the ion channel proteins, here we developed membrane-embedded synthetic chiral receptors 1 and 2 with different terminal structures, where receptor 1 has hydrophobic triisopropylsilyl (TIPS) groups and receptor 2 has hydrophilic hydroxy groups. The receptors have ligand-binding units that interact with cationic amphiphiles such as 2-phenethylamine (PA). Conductance study revealed that the receptors hardly show ion transportation at the ligand-free state. After ligand binding involving a conformational change, receptor 1 bearing TIPS termini displays a significant current enhancement due to ion transportation. The current substantially diminishes upon addition of β-cyclodextrin (βCD) that scavenges the ligand from the receptor. Importantly, the receptor again turns into the conductive state by the second addition of PA, and the activation/deactivation of the ion transportation can be repeated. In contrast, receptor 2 bearing the hydroxy terminal groups hardly exhibits ion transportation, suggesting the importance of terminal TIPS groups of 1 that likely anchor the receptor in the membrane.

Micrometer-Size Vesicle Formation Triggered by UV Light

Vesicle formation is a fundamental kinetic process related to the vesicle budding and endocytosis in a cell. In the vesicle formation by artificial means, transformation of lamellar lipid aggregates into spherical architectures is a key process and known to be prompted by e.g. heat, infrared irradiation, and alternating electric field induction. Here we report UV-light-driven formation of vesicles from particles consisting of crumpled phospholipid multilayer membranes involving a photoactive amphiphilic compound composed of 1,4-bis(4-phenylethynyl)benzene (BPEB) units. The particles can readily be prepared from a mixture of these components, which is casted on the glass surface followed by addition of water under ultrasonic radiation. Interestingly, upon irradiation with UV light, micrometer-size vesicles were generated from the particles. Neither infrared light irradiation nor heating prompted the vesicle formation. Taking advantage of the benefits of light, we successfully demonstrated micrometer-scale spatiotemporal control of single vesicle formation. It is also revealed that the BPEB units in the amphiphile are essential for this phenomenon.

Single-cell E. coli response to an instantaneously applied chemotactic signal.

In response to an attractant or repellant, an Escherichia coli cell controls the rotational direction of its flagellar motor by a chemotaxis system. When an E. coli cell senses an attractant, a reduction in the intracellular concentration of a chemotaxis protein, phosphorylated CheY (CheY-P), induces counterclockwise (CCW) rotation of the flagellar motor, and this cellular response is thought to occur in several hundred milliseconds. Here, to measure the signaling process occurring inside a single E. coli cell, including the recognition of an attractant by a receptor cluster, the inactivation of histidine kinase CheA, and the diffusion of CheY and CheY-P molecules, we applied a serine stimulus by instantaneous photorelease from a caged compound and examined the cellular response at a temporal resolution of several hundred microseconds. We quantified the clockwise (CW) and CCW durations immediately after the photorelease of serine as the response time and the duration of the response, respectively. The results showed that the response time depended on the distance between the receptor and motor, indicating that the decreased CheY-P concentration induced by serine propagates through the cytoplasm from the receptor-kinase cluster toward the motor with a timing that is explained by the diffusion of CheY and CheY-P molecules. The response time included 240 ms for enzymatic reactions in addition to the time required for diffusion of the signaling molecule. The measured response time and duration of the response also revealed that the E. coli cell senses a similar serine concentration regardless of whether the serine concentration is increasing or decreasing. These detailed quantitative findings increase our understanding of the signal transduction process that occurs inside cells during bacterial chemotaxis.

Chromatography-free synthesis of monodisperse oligo(ethylene glycol) mono-p-toluenesulfonates and quantitative analysis of oligomer purity

Poly(ethylene glycol) (PEG) is a common building block for complex functional molecules. Because of its chain flexibility and hydrophilicity, it is widely used in bioconjugation chemistry. Modern applications require PEGs of high purity, hence well-defined, monodisperse PEG oligomers are gaining more attention. Here we report a large-scale synthetic procedure for monodisperse oligo(ethylene glycol) mono-p-toluenesulfonates, convenient intermediates of various heterobifunctional PEG derivatives. This method features no chromatographic purification, low total material cost and high purity of the products. In addition, oligomer dispersity of the products is determined quantitatively using reverse-phase HPLC and a number of bioconjugation-related functionalized PEG derivatives are prepared from the synthesized toluenesulfonates.