Minako Kondo

Reactive Dynamics in Confined Liquids : Ultrafast Torsional Dynamics of Auramine O in Nanoconfined Water in Aerosol OT Reverse Micelles

The effects of confinement on the ultrafast torsional reaction of auramine O in aqueous solution are investigated through ultrafast fluorescence up-conversion with 50 fs time resolution. The aqueous solution is confined in nanoscale water droplets by an ionic surfactant. The torsional motion is orders of magnitude slower in the confined droplets than in bulk aqueous solution. The dynamics become faster with increasing radius of the nanodroplet but never reach the bulk value, even when the radius is as large as 10 nm. Time-dependent fluorescence spectra were constructed and subsequently analyzed using a one-dimensional generalized Smoluchowski equation. An accurate description of the data was achieved using a time-dependent diffusion coefficient. This is suggested to arise because the medium friction reflects dynamics on a broad range of time scales spanning the reaction dynamics. The friction recovered suggests strongly hindered motion in the confined droplet and can be qualitatively related to solvation dynamics measured in AOT, consistent with auramine O torsional dynamics being accompanied by intramolecular charge redistribution.

Chemically modulating the photophysics of the GFP chromophore.

There is growing interest in engineering the properties of fluorescent proteins through modifications to the chromophore structure utilizing mutagenesis with either natural or unnatural amino acids. This entails an understanding of the photophysical and photochemical properties of the modified chromophore. In this work, a range of GFP chromophores with different alkyl substituents are synthesized and their electronic spectra, pH dependence, and ultrafast fluorescence decay kinetics are investigated. The weakly electron donating character of the alkyl substituents leads to dramatic red shifts in the electronic spectra of the anions, which are accompanied by increased fluorescence decay times. This high sensitivity of electronic structure to substitution is also characteristic of some fluorescent proteins. The solvent viscosity dependence of the decay kinetics are investigated, and found to be consistent with a bimodal radiationless relaxation coordinate. Some substituents are shown to distort the planar structure of the chromophore, which results in a blue shift in the electronic spectra and a strong enhancement of the radiationless decay. The significance of these data for the rational design of novel fluorescent proteins is discussed.

Ultrafast dynamics of protein proton transfer on short hydrogen bond potential energy surfaces: S65T/H148D GFP.

Ultrafast proton transfer dynamics on a short H-bond in a protein were studied through the time-resolved fluorescence of the S65T/H148D green fluorescent protein (GFP) mutant. In response to the change in chromophore pK(a) upon excitation, the donor-proton-acceptor structure evolves on a sub-100 fs time scale, followed by picosecond time scale vibrational cooling and host structure reorganization.

Reactive dynamics in micelles: auramine O in solution and adsorbed on regular micelles.

The role of confinement in the suppression of the excited-state reaction of the dye molecule auramine O in nanoscale water droplets is investigated by contrasting the behavior of the dye in solution and on regular and reverse micelles. Auramine O photophysics are studied in bulk water, at the interface between regular micelles and bulk water, and in the aerosol OT (AOT)-stabilized aqueous nanodroplet. It is shown that the reaction of auramine O in bulk water is to a first approximation determined by aqueous solvation dynamics rather than solvent viscosity. This is in contrast to the result in more viscous and slowly relaxing solvents, where the solvent viscosity controls the rate. This result suggests the possibility of multiple reaction pathways on the excited-state surface. The reaction rate at the regular micelle-water interface is slower than in bulk water but significantly faster than in nanoconfined water, indicating a distinct effect of confinement on the reaction rate. It is suggested that the degree of perturbation of the water structure at the interface is the factor controlling the rate of the reaction. Specifically, the water structure is strongly perturbed at the AOT-confined water interface, suppressing the ability of fast collective solvent reorientation to promote the auramine O excited-state reaction. This effect is less marked at the interface between the micelle and bulk water. The contrast between these results indicates a route whereby confinement modifies ultrafast reaction dynamics in micelles.

Reactive Dynamics in Confined Liquids: Interfacial Charge Effects on Ultrafast Torsional Dynamics in Water Nanodroplets

The excited-state dynamics of a reactive dye molecule, auramine O, have been studied in nanoscale water droplets stabilized by a nonionic surfactant. Spectral dynamics were measured as a function of the radius of the water nanodroplet with 50 fs time resolution using time-resolved fluorescence up-conversion method. Qualitatively, the effect of confinement is to dramatically slow the rate of the reaction compared to that of bulk water. Data were quantitatively analyzed using the one-dimensional generalized Smoluchowski equation assuming a time-dependent diffusion coefficient. The results were contrasted with our earlier analysis of auramine O in aqueous nanodroplets stabilized by the ionic surfactant AOT. The excited-state reaction is slower in the nonionic surfactant, showing that interfacial charge is not required to suppress reactions in nanoscale water droplets. The location of the dye in the heterogeneous micelle is investigated by comparing the absorption spectra of AO in the micelle with those of a water- polyethyleneglycol mixture (to mimic the surfactant head group). The results suggest that the charged dye is located in the water phase.

Ultrafast Infrared Spectroscopy of an Isotope-Labeled Photoactivatable Flavoprotein

The blue light using flavin (BLUF) domain photosensors, such as the transcriptional antirepressor AppA, utilize a noncovalently bound flavin as the chromophore for photoreception. Since the isoalloxazine ring of the chromophore is unable to undergo large-scale structural change upon light absorption, there is intense interest in understanding how the BLUF protein matrix senses and responds to flavin photoexcitation. Light absorption is proposed to result in alterations in the hydrogen-bonding network that surrounds the flavin chromophore on an ultrafast time scale, and the structural changes caused by photoexcitation are being probed by vibrational spectroscopy. Here we report ultrafast time-resolved infrared spectra of the AppA BLUF domain (AppA(BLUF)) reconstituted with isotopically labeled riboflavin (Rf) and flavin adenine dinucleotide (FAD), which permit the first unambiguous assignment of ground and excited state modes arising directly from the flavin carbonyl groups. Studies of model compounds and DFT calculations of the ground state vibrational spectra reveal the sensitivity of these modes to their environment, indicating that they can be used as probes of structural dynamics.

Ultrafast reaction dynamics in nanoscale water droplets confined by ionic surfactants

The excited state dynamics of the dye molecule auramine O have been measured in nanoscale size water droplets stabilized by ionic surfactants using ultrafast fluorescence up-conversion. Specifically, the reaction dynamics have been measured as a function of the size of the water droplet in the range 1–10 nm and as a function of the counterion charge. The data are analysed quantitatively using a generalized Smoluchowski equation. The rate of the reaction in the reverse micelle is dramatically decreased compared to the bulk water environment. It increases as nanodroplet size increases, but never attains the bulk value. The data are not well described by a ‘core-shell’ model, i.e. by assuming a slow reaction in the interface (shell) and a bulk-like decay in the core. The effect of changing the counterion is small, with three different singly charged counterions showing essentially identical reaction dynamics, while the double charged Ca2+ counterion only slightly slows the reaction. This insensitivity to counterion is in contrast to the observations of solvation dynamics experiments and molecular dynamics simulation. The origins of these differences are discussed in terms of the solute reaction coordinate being more sensitive to different motions of the confined aqueous solvent than solvation dynamics and to more delocalised properties of the dispersed phase than are detected in simulations.

Characterization of trans-2-[4-[(dimethylamino)styryl]benzothiazole as an ultrafast isomerization probe and a modifed Kramers theory analysis.

The photophysics of trans-2-[4-(dimethylamino)styryl]benzothiazole (DMASBT) was investigated by electronic structure calculations and steady-state and time-resolved emission spectroscopy in a wide range of solvents, including temperature and pressure dependence. DMASBT undergoes a facile photoinduced trans-cis isomerization similar to the reaction of trans-stilbene. The cis isomer has a lifetime of ∼1 ps and does not contribute appreciably to the steady-state emission spectrum. The absorption spectrum of the cis form overlaps that of the trans form such that considerable care is needed in determining correct emission quantum yields. The approximate equality of absorption and emission transition moments of DMASBT in all solvents indicates that absorption and emission involve a single excited state with high radiative rate. The low quantum yields of DMASBT in low-viscosity solvents reflect emission lifetimes in the 20-50 ps range. The nonradiative rates of DMASBT, which are assumed to measure the rate of isomerization in S1, depend upon both solvent viscosity and polarity. A modified Kramers analysis, which allows for a polarity-dependent barrier height, provides a satisfactory description of these rates but only if it is assumed that friction in alcohol solvents is more weakly dependent upon viscosity than in other types of solvents.

Reactive Dynamics in Nanoscale Water droplets Confined in Inverse Micelles

Ultrafast excited state reactions of Auramine are studied in inverse micelles with water droplets between 1 and 10nm. Dynamics, inhomogeneous and a function of droplet size, are discussed in terms of interfacial and confinement effects.

Ultrafast Vibrational Dynamics in the AppA Blue Light Sensing Protein

The mechanism of blue light sensing in the photoactive protein AppA is investigated by transient infra-red spectroscopy. Modes associated with the flavin excited state and perturbation of the protein are detected.