MPAI (mass probes aided ionization) method for total analysis of biomolecules by mass spectrometry.

We have designed and synthesized various mass probes, which enable us to effectively ionize various molecules to be detected with mass spectrometry. We call the ionization method using mass probes the "MPAI (mass probes aided ionization)" method. We aim at the sensitive detection of various biological molecules, and also the detection of bio-molecules by a single mass spectrometry serially without changing the mechanical settings. Here, we review mass probes for small molecules with various functional groups and mass probes for proteins. Further, we introduce newly developed mass probes for proteins for highly sensitive detection.     

Development of 14-epi-19-nortachysterol and its unprecedented binding configuration for the human vitamin D receptor

In the study of the synthesis of 14-epi-19-norprevitamin D(3), we found 14-epi-19-nortachysterol derivatives through C6,7-cis/trans isomerization. We also succeeded in their chemical synthesis and revealed their marked stability and potent VDR binding affinity. To the best of our knowledge, this is the first isolation of stable tachysterol analogues. Surprisingly, 14-epi-19-nortachysterol derivatives exhibited an unprecedented binding configurations for the ligand binding pocket in hVDR, C5,6-s-trans and C7,8-s-trans triene configurations, which were opposite the natural C7,8-ene-configuration of 1α,25(OH)(2)D(3).     

Enantioselective cyclization of 4-alkenoic acids via an oxidative allylic C-H esterification

An enantioselective intramolecular oxidative cyclization of 4-alkenoic acids was developed. The reaction proceeded via a π-allyl Pd intermediate generated by an allylic C-H activation to give γ-lactone derivatives with moderate to good enantioselectivity. Spiro bis(isoxazoline) ligand, SPRIX, was indispensable for this asymmetric transformation.     

Statistical mechanics of protein allostery: roles of backbone and side-chain structural fluctuations

A statistical mechanical model of allosteric transition of proteins is developed by extending the structure-based model of protein folding to cases that a protein has two different native conformations. Partition function is calculated exactly within the model and free-energy surfaces associated with allostery are derived. In this paper, the model of allosteric transition proposed in a previous paper [Proc. Natl. Acad. Sci. U.S.A 134, 7775 (2010)] is reformulated to describe both fluctuation in side-chain configurations and that in backbone structures in a balanced way. The model is applied to example proteins, Ras, calmodulin, and CheY: Ras undergoes the allosteric transition between guanosine diphosphate (GDP)-bound and guanosine triphosphate (GTP)-bound forms, and the model results show that the GDP-bound form is stabilized enough to prevent unnecessary signal transmission, but the conformation in the GTP-bound state bears large fluctuation in side-chain configurations, which may help to bind multiple target proteins for multiple pathways of signaling. The calculated results of calmodulin show the scenario of sequential ordering in Ca(2+) binding and the associated allosteric conformational change, which are realized though the sequential appearing of pre-existing structural fluctuations, i.e., fluctuations to show structures suitable to bind Ca(2+) before its binding. Here, the pre-existing fluctuations to accept the second and third Ca(2+) ions are dominated by the side-chain fluctuation. In CheY, the calculated side-chain fluctuation of Tyr106 is coordinated with the backbone structural change in the β4-α4 loop, which explains the pre-existing Y-T coupling process in this protein. Ability of the model to explain allosteric transitions of example proteins supports the view that the large entropic effects lower the free-energy barrier of allosteric transition.     

Synthesis of π-expanded O-chelated boron-dipyrromethene as an NIR dye

A π-expanded BODIPY dye with an intramolecular boronate skeleton was synthesized by retro Diels-Alder reaction of the bicyclo[2.2.2]octadiene (BCOD)-fused BODIPY and the subsequent O-chelation. The photophysical and electrochemical properties were examined by UV-vis, fluorescence, CV measurements, and DFT calculation. This BODIPY exhibited the absorption and emission over a visible-NIR region at 600-850 nm. O-chelated BODIPY showed a bathocromic shift compared to F-BODIPYs. This dye showed a bright fluorescence emission at 733 nm with the high Φ value of 0.58.     

Extracellular Production and Characterization of <Emphasis Type="BoldItalic">Streptomyces</Emphasis> X-prolyl Dipeptidyl Aminopeptidase

X-prolyl dipeptidyl aminopeptidases (X-PDAPs) are useful in various food industries. In this study, we performed sequence-based screening to obtain a stable X-PDAP enzyme from thermophilic Streptomyces strains. We found three genes that encoded X-PDAP from Streptomyces thermoluteus subsp. fuscus NBRC 14270 (14270 X-PDAP), Streptomyces thermocyaneoviolaceus NBRC 14271 (14271 X-PDAP), and Streptomyces thermocoerulescens NBRC 14273, which were subsequently cloned and sequenced. The deduced amino acid sequences of these genes showed high similarity, with ~80% identity with each other. The isolated X-PDAPs and an X-PDAP from Streptomyces coelicolor were expressed in Streptomyces lividans using a hyperexpression vector: pTONA5a. Among these genes, only 14270 and 14271 X-PDAPs caused overexpression and extracellular production without artificial signal peptides. We also characterized the biochemical properties of purified 14271 X-PDAP. In addition, we found that, in peptide synthesis via an aminolysis reaction, this enzyme recognized d-amino acid derivatives as acyl acceptors, similar to l-amino acid derivatives.     

Characteristics of microwaves on second generation nitrogen-doped TiO2 nanoparticles and their effect on photoassisted processes

Characteristics of 2.45-GHz microwave radiation were examined on two second generation nitrogen-doped TiO₂ nanomaterials prepared by annealing Degussa P-25 TiO₂ and Ishihara ST-01 TiO₂ at 400 °C and 500 °C, respectively, in air in the presence of urea, and for comparison on the undoped pristine samples. Band gaps of all four samples were determined by diffuse reflectance spectroscopy. Both the sizes and the BET specific surface areas of the TiO₂ particles were determined, together with dielectric constants and dielectric loss factors. Nitrogen doping caused the size to increase and surface area to decrease. Temperature–time profiles showed that the heating efficiency of the N-doped specimens by the microwaves was greater, particularly significant for the N-doped P25 sample, but rather small for the N-ST01 sample. The effect of microwaves on the surface optical phonons of the samples, with and without UV–vis irradiation, was examined by an in situ Raman spectroscopic technique; for the undoped P-25 and nitrogen-doped N-P25 TiO₂ systems the effect was negligible. By contrast, microwave irradiation of Ishihara ST-01 and nitrogen-doped N-ST01 TiO₂ samples showed significant changes in the 144-cm^?1 optical phonons. Results infer a microwave thermal effect on the Ishihara ST-01 and N-ST01 specimens, whereas for the Degussa P-25 samples the microwaves also imparted a specific effect as the microwaves influenced the N-dopant sites in contrast to the ST-01 systems where the dopant sites were unaffected as evidenced by temperature–time profiles. The microwave-/photo-assisted degradation of 4-chlorophenol under various conditions of UV–vis irradiation and conventional heating, as opposed to microwave heating, confirms the specific microwave effect for the P-25 systems.     

Enzymatic Production of Ferulic Acid from Defatted Rice Bran by Using a Combination of Bacterial Enzymes

Ferulic acid (FA), which is present in the cell walls of some plants, is best known for its antioxidant property. By combining a commercial enzyme that shows FA esterase activity with several Streptomyces carbohydrate-hydrolyzing enzymes, we succeeded in enhancing the enzymatic production of FA from defatted rice bran. In particular, the combination of three xylanases, an α-l-arabinofuranosidase, and an acetyl xylan esterase from Streptomyces spp. produced the highest increase in the amount of released FAs among all the enzymes in the Streptomyces enzymes library. This enzyme combination also had an effect on FA production from other biomasses, such as raw rice bran, wheat bran, and corncob.     

Synthetic Approach Toward Antibiotic Tunicamycins - VII Synthesis of Tunicamine and Tunicaminyl Uracil Derivative

A higher carbon carbohydrate moiety of antibiotic tunica-mycins named tunicamine has been synthesized in a protected form. The key reaction step of the synthesis is a potassium fluoride catalyzed Henry reaction of a 5-nitroribose derivative and a dial do-galactosamine derivative. The tunicamine derivative has been converted to a tunicaminyl uracil derivative by condenation with bis(trimethylsilyl) uracil.     

Synthesis and Application of Tailored Graft Copolymers from Polystyrene Macromonomer

Comb graft copolymers prepared by macromonomer technique has become applicable for permanent modification of polymer surfaces. In this paper the association and adsorption behavior in solution of the well designed graft copolymer prepared from polystyrene macromonomer and various hydrophilic comonomer was studied and the mechanism of surface accumulation of hydrophilic segment was clarified by surface analysis of polystyrene containing the graft copolymers. Wetting of modified polymer surfaces and the permeability of gases through the film was also discussed.