Browsing by Author "Hiramatsu, Hirotsugu"

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A Vertical Flow Method for Sensitive Raman Protein Measurement in Aqueous Solutions
(2019-08-06)
Li, Shu-Chi
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Hiramatsu, Hirotsugu
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交大名義發表
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應用化學系
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應用化學系分子科學碩博班
Raman scattering is intrinsically faint. Raman spectroscopy would be more valuable with improvements in the signal detection efficiency. To improve the signal detection efficiency, we propose a vertical flow method, which is a derivative of the liquid core optical fiber technique employed for sensitive Raman signal detection. In the vertical flow method, the sample solution flows from a pinhole to prepare a liquid column wrapped with air and uses total reflection at the sample-air interface to confine the excitation beam and Raman signal. The Raman signal emerges from the pinhole for efficient collection. This method enhanced Raman signal intensity by up to 90 times. When measuring a bovine serum albumin aqueous solution, the limit of detection (LOD) was 0.029 +/- 0.003 mg mL(-1) (0.44 +/- 0.05 mu M). We discuss the signal enhancement factor dependence on several parameters in the vertical flow method. Our method provides a simple technique to improve Raman spectroscopy sensitivity using universal materials.
Application of IR spectra of two successive isotope labeled residues to the evaluation of dihedral angles of polyproline II structure
(2019-03-01)
Miyata, Daisuke
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Hiramatsu, Hirotsugu
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Nakabayashi, Takakazu
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交大名義發表
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應用化學系
The method to evaluate the dihedral angles of a polypeptide using IR absorption spectroscopy was applied to a polyproline type II (PPII) structure. We have prepared (1)PPPPPPG(7)-NH2 with the two successive residues labeled by C-13=O-1(8). The doublet of the C-13=O-1(8) stretching band due to the two successive labeled residues was observed, which comes from the vibrational coupling between the labeled residues. The differences in the peak wavenumber and the intensity of the doublet band were analyzed using the GF matrix method and ab initio molecular orbital calculations. The obtained dihedral angles of the labeled residues were compared with the reported values.
Citrullination and deamidation affect aggregation properties of amyloid -proteins
(2016-12)
Osaki, Dai
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Hiramatsu, Hirotsugu
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應用化學系
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Department of Applied Chemistry
Citrullination and deamidation, which are aging-related posttranslational modifications, increase the number of negative charges on amyloid -protein (A) at neutral pH. We investigated the effects of these modifications on the fibrillation properties of A. The Arg5Cit modification of A(1-40) did not affect the fibrillation rate, and brought -sheet structures unlike that in the A(1-40) fibril. The Asn27Asp modification of A(1-40) stopped the fibrillation and induced the formation of aggregates that involved an anti-parallel -sheet. A(1-42) with the Arg5Cit modification showed increased solubility in aqueous media, and its fibril formation became slower than that of A(1-42). The modification did not change the parallel -sheet structure of the fibrils. A(1-42) with the Asn27Asp modification partially formed fibrils that involved the parallel -sheet structure. Using the thioflavin T (ThT) assay, an increased fraction of the soluble oligomer of each A analog was transiently detected during fibrillation. An increase in the number of negative charges at basic pH affected the aggregation properties of A in a manner different from that with the modifications, suggesting that change in properties of the posttanslationally modified residues rather than the number of charges in the peptide was important.
Directly Probing Intermolecular Structural Change of a Core Fragment of beta(2)-Microglobulin Amyloid Fibrils with Low-Frequency Raman Spectroscopy
(2017-01-26)
Shigeto, Shinsuke
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Chang, Chun-Fu
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Hiramatsu, Hirotsugu
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應用化學系
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Department of Applied Chemistry
Amyloid fibrils, which are ordered aggregates of proteins or peptides, have attracted keen interest because their deposition causes serious human diseases. Despite many studies utilizing X-ray crystallography, solid-state NMR, and other methods, intermolecular interactions governing the fibril formation remain largely unclear. Here, we used low-frequency Raman (LFR) spectroscopy to investigate the intermolecular beta-sheet structure of a core fragment of beta(2)-microglobulin amyloid fibrils, beta(2)m(21-29), in aqueous buffer solutions. The LFR spectra (approximately 10-200 cm(-1)) of beta(2)m(21-29) amyloid fibrils measured at different pH values (ranging from 6.8 to 8.0) revealed a broad-spectral pattern with a maximum at similar to 80 cm(-1) below pH 7.2 and at similar to 110 cm(-1) above pH 7.4. This observation is attributed to a pH-dependent structural change from an antiparallel to a parallel intermolecular beta-sheet structure. Multivariate curve resolution-alternating least-squares (MCR-ALS) analysis enabled us to decompose the apparently monotonous LFR spectra into three distinctly different contributions: intermolecular vibrations of the parallel and antiparallel beta-sheets and intramolecular vibrations of the peptide backbone. Peak positions of the obtained LFR bands not only exhibit a much more pronounced difference between the two beta-sheets than the conventional amide I band, but they also suggest stronger intermolecular interaction, due presumably to the hydrophobic effect, in the parallel beta-sheet than in the antiparallel beta-sheet. The present results show that LFR spectroscopy in combination with the MCR-ALS analysis holds promise for real-time tracking of the intermolecular dynamics of amyloid fibril formation under physiological conditions.
Electric field effects on 1-hydroxyacetone revealed by IR electroabsorption spectroscopy
(2019-01-16)
Hiramatsu, Hirotsugu
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交大名義發表
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應用化學系
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應用化學系分子科學碩博班
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National Chiao Tung University
Electric field effect was investigated on 1-hydroxyacetone (HA) as a model compound of the acyclic form of saccharide moiety employing infrared electroabsorption spectroscopy. We examined the structural tautomerization of the hydroxyacetyl group of HA in detail by monitoring change in IR absorption intensity of the C=O stretching band in the range of 1650-1850 cm(-1). The observed changes in absorbance were analyzed by considering a change in the equilibrium of HA dimer, HA monomer, endiol, and enol forms, as well as the orientational polarization of each tautomer. Conversion from the HA dimer to the enol was detected.
Evaluation of Dihedral Angles of Peptides Using IR Bands of Two Successive Isotope Labeled Residues
(2019-01-01)
Okabe, Hitomi
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Miyata, Daisuke
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Nakabayashi, Takakazu
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Hiramatsu, Hirotsugu
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交大名義發表
The infrared (IR) absorption bands due to peptide bonds (amide bands) have long been used to determine the secondary structure of a peptide and to analyze intra- and intermolecular interactions between amides. In the present study, the dihedral angles of a residue in peptides have also been evaluated using the amide I IR band of two successive residues with isotope labeling. The two successive residues labeled with the C-13 and O-18 isotopes give the doublet amide I IR band and the intensity ratio (R-int) and the difference in peak position (Delta nu) of the doublet band were analyzed using GF matrix and ab initio molecular orbital calculations. We obtained the two-dimensional calculation maps of R-int and Delta nu against the two dihedral angles. The crossing point of the curves of R-int and Delta nu is the two dihedral angles of the measured residue. The evaluated dihedral angles of the simple peptides are compared with the reported values. We discuss the limitation and the application of the present method to biopolymers from the obtained results.
Generation of self-clusters of galectin-1 in the farnesyl-bound form
(2016-09-14)
Yamaguchi, Kazumi
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Niwa, Yusuke
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Nakabayashi, Takakazu
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Hiramatsu, Hirotsugu
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應用化學系
Ras protein is involved in a signal transduction cascade in cell growth, and cluster formation of H-Ras and human galectin-1 (Gal-1) complex is considered to be crucial to achieve its physiological roles. It is considered that the complex is formed through interactions between Gal-1 and the farnesyl group (farnesyl-dependent model), post-translationally modified to the C-terminal Cys, of H-Ras. We investigated the role of farnesyl-bound Gal-1 in the cluster formation by analyzing the structure and properties of Gal-1 bound to farnesyl thiosalicylic acid (FTS), a competitive inhibitor of the binding of H-Ras to Gal-1. Gal-1 exhibited self-cluster formation upon interaction with FTS, and small-and large-size clusters were formed depending on FTS concentration. The galactoside-binding pocket of Gal-1 in the FTS-bound form was found to play an important role in small-size cluster formation. Large-size clusters were likely formed by the interaction among the hydrophobic sites of Gal-1 in the FTS-bound form. The present results indicate that Gal-1 in the FTS-bound form has the ability to form self-clusters as well as intrinsic lectin activity. Relevance of the self-clustering of FTS-bound Gal-1 to the cluster formation of the H-Ras-Gal-1complex was discussed by taking account of the farnesyl-dependent model and another (Raf-dependent) model.
Linear, mixed-valent homocatenated tri-tin complexes featuring Sn-Sn bonds
(2020-06-25)
Chang, Wei-Chieh
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Raj, Ankit
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Hiramatsu, Hirotsugu
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Li, Han-Jung
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Ziegler, Micah S.
A series of tri-tin complexes((LSn)-Sn-Ph)(3)Xwith triple-decker structures (L-Ph= 2,5-di(o-pyridyl)-3,4-diphenylpyrrolate; X = Cl, AlCl4, OTf, and PF6) was synthesized by reducingL(Ph)SnClwith (LiBBu3H)-Bu-s and subsequent reactions. Structural characterization of((LSn)-Sn-Ph)(3)Clrevealed a Sn-Sn-Sn core, and DFT calculations suggest that its HOMO is primarily sigma-bonding along the tri-tin framework.((LSn)-Sn-Ph)(3)Clreacts with W(CO)(5)THF to afford((LSn)-Sn-Ph)(2)(W(CO)(5))(2)andL(Ph)SnCl, implying that((LSn)-Sn-Ph)(3)Clmay exhibit dynamic behavior in solution.
Preparation and photo-induced activities of water-soluble amyloid beta-C-60 complexes
(2018-01-01)
Hasunuma, Naoki
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Kawakami, Masahiro
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Hiramatsu, Hirotsugu
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Nakabayashi, Takakazu
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應用化學系
We have shown that fullerene (C-60) becomes soluble in water by mixing fullerene and amyloid beta peptide (A beta(40)) whose fibril structures are considered to be associated with Alzheimer's disease. The water-solubility of fullerene arises from the generation of a nanosized complex between fullerene and the monomer species of A beta(40) (A beta(40)-C-60). The prepared A beta(40)-C-60 exhibits photo-induced activity with visible light to induce the inhibition of A beta(40) fibrillation and the cytotoxicity for cultured HeLa cells. The observed photo-induced phenomena result from the generation of singlet oxygen via photoexcitation, inducing oxidative damage to A beta(40) and HeLa cells. The oxidized A beta(40) following photoexcitation of A beta(40)-C-60 was confirmed by mass spectrometry.
Tautomer Structures in Ketose-Aldose Transformation of 1,3-Dihydroxyacetone Studied by Infrared Electroabsorption Spectroscopy
(2019-12-19)
Chen, Szu-Hua
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Hiramatsu, Hirotsugu
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交大名義發表
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應用化學系
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應用化學系分子科學碩博班
The acyclic form of monosaccharides exists in a structural equilibrium, with aldose having the aldehyde group and ketose the ketone group (ketose-aldose equilibrium). A basic catalyst facilitates their transformation, which affects the chemical properties of the monosaccharide. In this study, we investigated the ketose-aldose transformation of 1,3-dihydroxyacetone (1,3-DHA), one of the simplest systems of the ketose-aldose equilibrium. We examined the effects of piperidine as the basic catalyst and used IR electroabsorption spectroscopy to study the responses to an external electric field. We analyzed the changes in IR absorption by considering the changes in the molecular orientation and number of molecules in response to the external electric field. The results of the analysis revealed the permanent dipole moment mu(P), an angle eta between mu(P) and mu(T) (the transition moment of the molecular vibration), and the equilibrium constants. The ketose-aldose transformation of 1,3-DHA can be explained in terms of the equilibrium of three states. In the presence of piperidine, a five-state equilibrium was concluded. On the basis of the experimental data, we propose plausible models of dihydroxyacetone, E-enediols, Z-enediol, or glyceraldehyde for each state. The results of our structural analysis of these tautomers provide a detailed understanding of the ketose-aldose transformation of acyclic saccharides and the effects of the basic catalyst.
The 532-nm-excited hyper-Raman spectroscopy of globular protein and aromatic amino acids
(1970-01-01)
Wen, Chun-, I
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Hiramatsu, Hirotsugu
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交大名義發表
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應用化學系
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應用化學系分子科學碩博班
Hyper-Raman (HR) spectroscopy reveals unique information about molecular structures, because of its different selection rule, compared with that of the infrared (IR) and Raman spectroscopy. The application of HR spectroscopy on biomolecules' research has been expected for a long time. Herein, we report the HR spectra of the amino acids tryptophan (Trp), tyrosine (Tyr), histidine, and bovine serum albumin (BSA) in aqueous solution, obtained at an excitation wavelength of 532 nm. The HR bands of BSA were mostly assigned to those of the aromatic amino acids. The predominance of the aromatic amino acids was explained by considering the resonance effect of the excitation beam to the electronic absorption around 266 nm, the double frequency of 532 nm (e.g. L-a of Trp, L-b of Tyr). Besides the Raman bands observed in the 266-nm-excited ultraviolet resonance Raman spectrum, some of the IR-active bands of Tyr were also detected. Hence, the HR spectroscopy serves as a new tool toward the investigation of proteins as well as of biomolecules and more complicated biological structures with the 532-nm excitation.
The EMBO Workshop Transducing Glycan Information into Function: Lessons from and for Galectins
(2017-03-01)
Hiramatsu, Hirotsugu
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交大名義發表
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National Chiao Tung University
Time-resolved FTIR study on the structural switching of human galectin-1 by light-induced disulfide bond formation
(2020-01-21)
Kuroi, Kunisato
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Kamijo, Mana
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Ueki, Mutsuki
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Niwa, Yusuke
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Hiramatsu, Hirotsugu
Disulfide bonds play a fundamental role in controlling the tertiary structure of proteins; the formation or cleavage of some disulfide bonds can switch the structures and/or functions of proteins. Human galectin-1 (hGal-1), which is a lectin protein, exemplifies how both structure and function are changed by disulfide bonds; the structure and sugar-binding ability of hGal-1 are altered by the formation and cleavage of its three intra-molecular disulfide bonds. In the present study, the dynamics of the structural change of hGal-1 by the formation of disulfide bonds were investigated by time-resolved FTIR spectroscopy combined with a modification in which its thiol groups (-SH) were replaced with S-nitrosylated groups (SNO). Photodissociation of NO from SNO in reduced hGal-1 induced disulfide bond formation and transformed it into the oxidised form. The structural change to the oxidised form involved three distinct kinetics with fast (<300 s), middle (similar to 600 s), and slow (similar to 6400 s) lifetimes. In an examination of hGal-1 in the lactose-bound form, structural changes owing to the release of substrate lactose were also observed upon disulfide bond formation. The present method using the photodissociation of NO is useful for monitoring the dynamics of structural changes following disulfide formation.