Koichi Tsukiyama

Relationship between the time, yield, and enantiomeric excess of asymmetric autocatalysis of chiral 2-alkynyl-5-pyrimidyl alkanol with amplification of enantiomeric excess

Experimental and kinetic analysis of asymmetric autocatalysis with amplification of ee in the enantioselective addition of diisopropylzinc to 2-alkynylpyrimidine-5-carbaldehyde using chiral 2-alkynyl-5-pyrimidyl alkanol with low ees are described.

Revised conformational assignments and conformational evolution of tyrosine by laser desorption supersonic jet laser spectroscopy

The number of conformers and their structures of tyrosine are reassigned on the basis of resonance enhanced multiphoton ionization (REMPI), ultraviolet-ultraviolet hole burning (UV-UV HB), infrared (IR) dip spectra, and quantum chemical calculations. From comparison between REMPI and UV-UV HB spectra, it was found that 12 conformers coexist in the supersonic jet. The structures of these conformers are determined by the IR spectra and theoretical calculations. The number of conformers is more than that reported in the previous reports (8 conformers), and is rationalized by the systematic formation of conformers from simpler molecules without substituents, just like evolution. The importance of dipole-dipole interaction between an amino-acid chain and hydroxyl group at the benzene ring was also discussed.

Effect of Mid-infrared Free-Electron Laser Irradiation on Refolding of Amyloid-Like Fibrils of Lysozyme into Native Form

Aggregation of lysozyme in an acidic solution generates inactive amyloid-like fibrils, with a broad infrared peak appearing at 1,610–1,630xa0cm−1, characteristic of a β-sheet rich structure. We report here that spontaneous refolding of these fibrils in water could be promoted by mid-infrared free-electron laser (mid-IR FEL) irradiation targeting the amide bands. The Fourier transform infrared spectrum of the fibrils reflected a β-sheet content that was as low as that of the native structure, following FEL irradiation at 1,620xa0cm−1 (amide I band); both transmission-electron microscopy imaging and Congo Red assay results also demonstrated a reduced fibril structure, and the enzymatic activity of lysozyme fibrils recovered to 70–90xa0% of the native form. Both irradiations at 1,535xa0cm−1(amide II band) and 1,240xa0cm−1 (amide III band) were also more effective for the refolding of the fibrils than mere heating in the absence of FEL. On the contrary, either irradiation at 1,100 or 2,000xa0cm−1 afforded only about 60xa0% recovery of lysozyme activity. These results indicate that the specific FEL irradiation tuned to amide bands is efficient in refolding of lysozyme fibrils into native form.

Infrared photodissociation spectroscopy of protonated ammonia cluster ions, NH4+(NH3)n (n=5–8), by using infrared free electron laser

Infrared photodissociation action spectra of protonated ammonia cluster ions, NH(4) (+)(NH(3))(n) (n=5-8), were measured in the range of 1020-1210 cm(-1) by using a tunable infrared free electron laser. Analyses by the density functional theory (DFT) show that the spectral features observed can be assigned to the nu(2) vibrational mode of the NH(3) molecules in NH(4) (+)(NH(3))(n). Size dependence of the spectra supports structural models obtained by the DFT calculations, in which the NH(4) (+) ion is solvated by the four nearest-neighbor NH(3) molecules. For NH(4) (+)(NH(3))(5), the spectrum between 1000 and 1700 cm(-1) was measured. The nu(4) bands of the NH(3) molecules and the NH(4) (+) ion were found in the range of 1420-1700 cm(-1).

Mid-infrared free-electron laser tuned to the amide I band for converting insoluble amyloid-like protein fibrils into the soluble monomeric form

A mid-infrared free-electron laser (FEL) is operated as a pulsed and linearly polarized laser with tunable wavelengths within infrared region. Although the FEL can ablate soft tissues with minimum collateral damage in surgery, the potential of FEL for dissecting protein aggregates is not fully understood. Protein aggregates such as amyloid fibrils are in some cases involved in serious diseases. In our previous study, we showed that amyloid-like lysozyme fibrils could be disaggregated into the native form with FEL irradiation specifically tuned to the amide I band (1,620xa0cm−1). Here, we show further evidence for the FEL-mediated disaggregation of amyloid-like fibrils using insulin fibrils. Insulin fibrils were prepared in acidic solution and irradiated by the FEL, which was tuned to either 1,620 or 2,000xa0cm−1 prior to the experiment. The Fourier transform infrared spectroscopy (FT-IR) spectrum after irradiation with the FEL at 1,620xa0cm−1 indicated that the broad peak (1,630–1,660xa0cm−1) became almost a single peak (1,652xa0cm−1), and the β-sheet content was reduced to 25 from 40xa0% in the fibrils, while that following the irradiation at 2,000xa0cm−1 remained at 38xa0%. The Congo Red assay as well as transmission electron microscopy observation confirmed that the number of fibrils was reduced by FEL irradiation at the amide I band. Size-exclusion chromatography analysis indicated that the disaggregated form of fibrils was the monomeric form. These results confirm that FEL irradiation at the amide I band can dissect amyloid-like protein fibrils into the monomeric form in vitro.

Decay dynamics of the long-range H¯Σg+1 state of D2 and H2: Experiment and theory

We present accurate experimental measurements of the lifetimes of rovibrational levels of the long-range H¯Σg+1 state for both D2 and H2, obtained directly from the observation of the time-dependent decay of the fluorescence from these excited levels. These results improve upon and extend those of Reinhold et al. [J. Chem. Phys. 112, 10754 (2000)]. Several decay pathways are open to these levels including fluorescence, predissociation, and autoionization. We present theoretical results for each of these processes, each calculated using the simplest but still appropriate level of theory. In particular, the theoretical calculations provide a quantitative explanation of the dramatic vibrational dependence of the observed lifetimes, the isotope dependence of the lifetimes for levels well localized within the H¯ potential well and therefore not subject to significant tunneling, and an insight into the role of enhanced tunneling in autoionization. In these calculations each of the rovibrational levels of the H¯...

Infrared photodissociation spectroscopy and density-functional calculations of protonated methanol cluster ions: Solvation structures of an excess proton.

Solvation structures of an excess proton in protonated methanol cluster ions, H(+)(CH(3)OH)(n) (n=5-8), were investigated by photodissociation spectroscopy in the middle infrared region (900-2300 cm(-1)) and by using density-functional theory. This work indicates that the excess proton is delocalized between two methanol molecules. Spectral features observed in the range 1400-1800 cm(-1) are attributed to vibrational modes involving collective motion of the shared proton and the two ligand molecules. At n=6-8, broad spectral features in the region above 1800 cm(-1) suggest coexistence of isomers in which the excess proton and a methanol molecule are tightly bound to form an ion core, CH(3)OH(2) (+).

Application of mid-infrared free-electron laser tuned to amide bands for dissociation of aggregate structure of protein

A mid-infrared free-electron laser (FEL) is a linearly polarized, high-peak powered pulse laser with tunable wavelength within the mid-infrared absorption region. It was recently found that pathogenic amyloid fibrils could be partially dissociated to the monomer form by the irradiation of the FEL targeting the amide I band (C=O stretching vibration), amide II band (N-H bending vibration) and amide III band (C-N stretching vibration). In this study, the irradiation effect of the FEL on keratin aggregate was tested as another model to demonstrate an applicability of the FEL for dissociation of protein aggregates. Synchrotron radiation infrared microscopy analysis showed that the α-helix content in the aggregate structure decreased to almost the same level as that in the monomer state after FEL irradiation tuned to 6.06u2005µm (amide I band). Both irradiations at 6.51u2005µm (amide II band) and 8.06u2005µm (amide III band) also decreased the content of the aggregate but to a lesser extent than for the irradiation at the amide I band. On the contrary, the irradiation tuned to 5.6u2005µm (non-absorbance region) changed little the secondary structure of the aggregate. Scanning-electron microscopy observation at the submicrometer order showed that the angular solid of the aggregate was converted to non-ordered fragments by the irradiation at each amide band, while the aggregate was hardly deformed by the irradiation at 5.6u2005µm. These results demonstrate that the amide-specific irradiation by the FEL was effective for dissociation of the protein aggregate to the monomer form.

Picosecond pulsed infrared laser tuned to amide I band dissociates polyglutamine fibrils in cells

Amyloid fibrils are causal substances for serious neurodegenerative disorders and amyloidosis. Among them, polyglutamine fibrils seen in multiple polyglutamine diseases are toxic to neurons. Although much efforts have been made to explore the treatments of polyglutamine diseases, there are no effective drugs to block progression of the diseases. We recently found that a free electron laser (FEL), which has an oscillation wavelength at the amide I band (Cu2009=u2009O stretch vibration mode) and picosecond pulse width, was effective for conversion of the fibril forms of insulin, lysozyme, and calcitonin peptide into their monomer forms. However, it is not known if that is also the case in polyglutamine fibrils in cells. We found in this study that the fibril-specific β-sheet conformation of polyglutamine peptide was converted into nonfibril form, as evidenced by the infrared microscopy and scanning-electron microscopy after the irradiation tuned to 6.08xa0μm. Furthermore, irradiation at this wavelength also changed polyglutamine fibrils to their nonfibril state in cultured cells, as shown by infrared mapping image of protein secondary structure. Notably, infrared thermography analysis showed that temperature increase of the cells during the irradiation was within 1xa0K, excluding thermal damage of cells. These results indicate that the picosecond pulsed infrared laser can safely reduce amyloid fibril structure to the nonfibril form even in cells.

Antimicrobial effect of blue light using Porphyromonas gingivalis pigment

The development of antibiotics cannot keep up with the speed of resistance acquired by microorganisms. Recently, the development of antimicrobial photodynamic therapy (aPDT) has been a necessary antimicrobial strategy against antibiotic resistance. Among the wide variety of bacteria found in the oral flora, Porphyromonas gingivalis (P. gingivalis) is one of the etiological agents of periodontal disease. aPDT has been studied for periodontal disease, but has risks of cytotoxicity to normal stained tissue. In this study, we performed aPDT using protoporphyrin IX (PpIX), an intracellular pigment of P. gingivalis, without an external photosensitizer. We confirmed singlet oxygen generation by PpIX in a blue-light irradiation intensity-dependent manner. We discovered that blue-light irradiation on P. gingivalis is potentially bactericidal. The sterilization mechanism seems to be oxidative DNA damage in bacterial cells. Although it is said that no resistant bacteria will emerge using aPDT, the conventional method relies on an added photosensitizer dye. PpIX in P. gingivalis is used in energy production, so aPDT applied to PpIX of P. gingivalis should limit the appearance of resistant bacteria. This approach not only has potential as an effective treatment for new periodontal diseases, but also offers potential antibacterial treatment for multiple drug resistant bacteria.