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Biomedical subjects

Nobuo Tanaka

Publications and source records attributed to Nobuo Tanaka.

16 recordsLinked to original sources

Structural insight into modest binding of a non-PXXP ligand to the signal transducing adaptor molecule-2 Src homology 3 domain.

Although some exceptional motifs have been identified, it is well known that the PXXP motif is the motif of ligand proteins generally recognized by the Src homology 3 (SH3) domain. SH3-ligand interactions are usually weak, with ordinary KD approximately 10 microM. The structural basis for a tight and specific association (KD = 0.24 microm) between Gads SH3 and a novel motif, PX(V/I)(D/N)RXXKP, was revealed in a previous structural analysis of the complex formed between them. In this paper, we report the crystal structure of the signal transducing adaptor molecule-2 (STAM2) SH3 domain in complex with a peptide with a novel motif derived from a ligand protein, UBPY. The derived KD value for this complex is 27 microM. The notable difference in affinity for these parallel complexes may be explained because the STAM2 SH3 structure does not provide a specificity pocket for binding, whereas the Gads SH3 structure does. Instead, the structure of STAM2 SH3 is analogous to that of Grb2 SH3 which, in addition to normal PXXP ligands, has also been shown to moderately recognize the novel motif discussed herein. Thus, the extremely tight interaction observed between Gads SH3 and the novel motif is caused not by an innate ability of the novel motif but rather by an evolutionary change in the Gads SH3 domain. Instead, SH3 domains of STAM2 and Grb2 retain the moderate characteristics of recognizing their ligand proteins like other SH3 domains for appropriate transient interactions between signaling molecules.

Adaptor Proteins, Signal Transducing↗

Crystallization and preliminary X-ray studies of xylanase 10B from Thermotoga maritima.

Xylanases catalyze the hydrolysis of the beta-1,4-glycosidic bonds of xylan, which is the second most abundant component of plant cell walls after cellulose. The recombinant xylanase 10B from Thermotoga maritima MSB8 was prepared and crystallized by the sitting-drop vapour-diffusion method using 40 mM zinc acetate, 20 mM MES buffer pH 6.0 and 3% ethanol. Intensity data were collected to 2.5 A resolution at beamline BL26B2 of SPring-8. Preliminary X-ray analysis showed that the crystal belongs to space group P2(1)2(1)2, with unit-cell parameters a = 77.3, b = 80.6, c = 58.2 A and one molecule per asymmetric unit.

Cloning, Molecular↗

Specific compositions of amyloid-beta peptides as the determinant of toxic beta-aggregation.

Alzheimer's disease (AD) may be caused by toxic aggregates formed from amyloid-beta (Abeta) peptides. By using Thioflavin T, a dye that specifically binds to beta-sheet structures, we found that highly toxic forms of Abeta-aggregates were formed at the initial stage of fibrillogenesis, which is consistent with recent reports on Abeta oligomers. Formation of such aggregates depends on factors that affect both nucleation and elongation. As reported previously, addition of Abeta42 systematically accelerated the nucleation of Abeta40, most likely because of the extra hydrophobic residues at the C terminus of Abeta42. At Abeta42-increased specific ratio (Abeta40: Abeta42 = 10: 1), on the other hand, not only accelerated nucleation but also induced elongation were observed, suggesting pathogenesis of early-onset AD. Because a larger proportion of Abeta40 than Abeta42 was still required for this phenomenon, we assumed that elongation does not depend only on hydrophobic interactions. Without any change in the C-terminal hydrophobic nature, elongation was effectively induced by mixing wild type Abeta40 with Italian variant Abeta40 (E22K) or Dutch variant (E22Q). We suggest that Abeta peptides in specific compositions that balance hydrophilic and hydrophobic interactions promote the formation of toxic beta-aggregates. These results may introduce a new therapeutic approach through the disruption of this balance.

Alzheimer Disease↗

On-column concentration of bisphenol A with one-step removal of humic acids in water.

An efficient extraction method for bisphenol A from environmental water including contaminants was developed using surface selective localization of functional group, on a polymeric separation device. The polymer utilized in this study was prepared through a kind of molecular imprinting technique, namely fragment imprinting effect utilizing a pseudo-template molecule (p-tert.-butylphenol) instead of bisphenol A. The concentration of bisphenol A onto the polymer device prepared, up to 1000 times concentration from environmental water including contaminants (humic acids), was achieved very easily with interesting exclusion effect for humic acids. The results obtained in this study suggest that molecular imprinting with the pseudo-template molecule is quite an effective way for selective concentration of the diluted target molecule from other contaminants including similar functional group with the target molecule.

Benzhydryl Compounds↗

A possible purification method of DNAs' fragments from humic matters in soil extracts using novel stimulus responsive polymer adsorbent.

Novel stimulus responsive, uniformly sized polymer-based adsorbent was prepared for a possible purification method of DNAs from humic matters in soil extracts. The prepared polymer adsorbent has a pair of anion exchangeable and cation exchangeable polymeric selectors, which are reversibly responded by the changes of column temperature as well as pH of mobile phase. At pH 5, DNAs and humic matters were completely adsorbed on the polymer adsorbent at room temperature, while up to 90% of the adsorbed DNAs were released and recovered at 70 degrees C with no release of adsorbed humic matters. Cleaning up of the polymer adsorbent could be performed by washing the adsorbed humic matters with alkaline mobile phase (pH 9) to recover those and realize repeatable use of the adsorbent.

Adsorption↗

Practical procedure for coma-free alignment using caustic figure.

The practical procedure for coma-free alignment using a single defocused transmission electron microscopy (TEM) image is presented. Caustic figures observed in the defocused TEM image of a focused probe are utilized. Coma-free alignment can be carried out by coinciding a bright-field spot with the center of a caustic curve as observed in an underfocus TEM image. With this method, beam tilt misalignment is reduced to the sub-mrad order (e.g. 0.3mrad for 300kV FEG-TEM). This can be done without intentional beam tilting, an amorphous specimen, high-resolution TEM images, or fast Fourier transform for diffractogram or cross-correlation, which are used in previous methods. Residual coma aberration is detected using the multiple Bragg images of a known crystal. Similarity between the present coma-free alignment and well-known STEM alignment using shadow image is discussed.

Journal Article↗

A tumour-associated DEAD-box protein, rck/p54 exhibits RNA unwinding activity toward c-myc RNAs in vitro.

BACKGROUND: The rck/p54 protein of 473 amino acids belongs to the family of DEAD-box/putative RNA helicase proteins. DEAD-box proteins have been implicated in a wide variety of cellular processes ranging from the initiation of protein synthesis and ribosome biosynthesis to premRNA splicing by means of modifying the RNA structure. Our previous data suggested that rck/p54 positively affected the translation initiation of c-myc mRNA. RESULTS: The data obtained from morphological studies and surface plasmon resonance assays clearly indicated that the protein specifically bound to c-myc RNA transcripts (RNAs) and exhibited RNA unwinding activity toward c-myc RNAs in the presence of ATP in vitro. Experiments using a deletion mutant of rck/p54 retaining only its N-terminal 289 amino acids demonstrated that the deleted C-terminal 184 amino acid domain is involved in the RNA unwinding activity. CONCLUSION: These findings strongly suggest that rck/p54 may play an important role in translation initiation by restructuring mRNAs even in the cell and contribute to carcinogenesis.

DEAD-box RNA Helicases↗

First observation of SiO2/Si(100) interfaces by spherical aberration-corrected high-resolution transmission electron microscopy.

SiO2/Si(100) interfaces were for the first time observed by a spherical aberration-corrected high-resolution transmission electron microscope in a cross-sectional mode. As the Fresnel fringes were not contrasted at the interfaces, the interfacial structures were clearly observed without the need for artificial image contrast. Atomic steps and defects on the Si(100) surfaces were accurately identified. Also, image simulations with the target imaging performance revealed oxygen atomic columns between silicon-silicon bonds. The present instrument is of potential use for semiconductor science and technology, even for the analysis of oxygen atoms at interfaces.

Journal Article↗

Novel surface modification of polymer-based separation media controlling separation selectivity, retentivity and generation of electroosmotic flow.

Uniformly sized packing materials based on synthetic polymer particles for high-performance liquid chromatography (HPLC) and capillary electrochromatography (CEC) have been prepared from polymerization mixtures containing methacrylic acid (MAA) as a functional monomer and by using a novel surface modification method. This "dispersion method" affords effectively modified separation media. Both the amount of MAA utilized in the preparation and reaction time affect the selectivity of chromatographic separation in both the HPLC and the CEC mode and electroosmotic flow. This detailed study revealed that the dispersion method effectively modified internal surface of macroporous separation media and, based on the amount of MAA introduced, exclusion mechanism for the separation of certain solutes could be observed.

Chromatography, High Pressure Liquid↗

Monolithic silica columns with various skeleton sizes and through-pore sizes for capillary liquid chromatography.

Reduction of through-pore size and skeleton size of a monolithic silica column was attempted to provide high separation efficiency in a short time. Monolithic silica columns were prepared to have various sizes of skeletons (approximately 1-2 microm) and through-pores (approximately 2-8 microm) in a fused-silica capillary (50-200 microm I.D.). The columns were evaluated in HPLC after derivatization to C18 phase. It was possible to prepare monolithic silica structures in capillaries of up to 200 microm I.D. from a mixture of tetramethoxysilane and methyltrimethoxysilane. As expected, a monolithic silica column with smaller domain size showed higher column efficiency and higher pressure drop. High external porosity (> 80%) and large through-pores resulted in high permeability (K = 8 x 10(-14) -1.3 x 10(-12) m2) that was 2-30 times higher than that of a column packed with 5-mirom silica particles. The monolithic silica columns prepared in capillaries produced a plate height of about 8-12 microm with an 80% aqueous acetonitrile mobile phase at a linear velocity of 1 mm/s. Separation impedance, E, was found to be as low as 100 under optimum conditions, a value about an order of magnitude lower than reported for conventional columns packed with 5-microm particles. Although a column with smaller domain size generally resulted in higher separation impedance and the lower total performance, the monolithic silica columns showed performance beyond the limit of conventional particle-packed columns under pressure-driven conditions.

Chromatography, Liquid↗

Monolithic silica columns for high-efficiency separations by high-performance liquid chromatography.

Generation of a large number of theoretical plates was attempted by capillary HPLC. Monolithic silica columns having small skeletons (ca. 2 microm) and large through-pores (ca. 8 microm) were prepared by a sol-gel method in a fused-silica capillary (50 microm I.D.), and derivatized to C18 phase by on-column reaction. High external porosity (>80%) and large through-pores resulted in high permeability (K= 1.2 x 10(-2) m2). The monolithic silica column in the capillary produced a plate height of about 12 microm in 80% acetonitrile at a linear velocity of 1 mm/s. Separation impedance, E value, was found to be as low as 200, that was about an order of magnitude lower than reported values for conventional columns packed with 5 microm particles. Reproducibility of preparation within +/- 15% was obtained for column efficiency and for pressure drop. It was possible to generate 100,000 plates by using a 130-cm column at very low pressure (<7 kg/cm2). A considerable decrease in column efficiency was observed at high linear velocity, and for solutes with large retention factors due to the slow mobile-phase mass transfer in the large through-pores. The monolithic silica columns, however, showed performance beyond the limit of conventional particle-packed columns in HPLC under favorable conditions.

Chromatography, High Pressure Liquid↗

Physically adsorbed chiral stationary phase of avidin on monolithic silica column for capillary electrochromatography and capillary liquid chromatography.

The physical adsorption method proposed previously has been successfully applied to a monolithic silica column. By virtue of the physical adsorption, a chiral stationary phase of avidin was prepared onto the silica monolith. The phase ratio of resulting stationary phase was evaluated with frontal analysis. The method proved to be comparable in phase ratio to the chemical bonding methods used in high-performance liquid chromatography (HPLC). Enantiomer separations were carried out in capillary electrochromatography (CEC) and capillary liquid chromatography (CLC) modes. Due to its larger phase ratio, the resulting column showed more powerful separation capability as compared to open-tubular CEC (OTCEC). Twelve chiral compounds were baseline-resolved. The resulting column showed high separation efficiency, with average theoretical plate numbers of 66 000/m for CLC and 122 000/m for CEC. Good reproducibility was observed, with RSD value less than 1.3% for retention time, retention factor and separation factor, and less than 6.6% for plate counts and resolution (n = 40). Fast separations were achieved with a short column. The test enantiomers were baseline-resolved within 4 min under CLC and CEC modes. In addition, field-enhanced sample injection (FESI) was coupled to CLC as well as CEC to improve the detection sensitivity.

Adsorption↗

The 2.0 A crystal structure of catalase-peroxidase from Haloarcula marismortui.

Catalase-peroxidase is a member of the class I peroxidase superfamily. The enzyme exhibits both catalase and peroxidase activities to remove the harmful peroxide molecule from the living cell. The 2.0 A crystal structure of the catalase-peroxidase from Haloarcula marismortui (HmCP) reveals that the enzyme is a dimer of two identical subunits. Each subunit is composed of two structurally homologous domains with a topology similar to that of class I peroxidase. The active site of HmCP is in the N-terminal domain. Although the arrangement of the catalytic residues and the cofactor heme b in the active site is virtually identical to that of class I peroxidases, the heme moiety is buried inside the domain, similar to that in a typical catalase. In the vicinity of the active site, novel covalent bonds are formed among the side chains of three residues, including that of a tryptophan on the distal side of the heme. Together with the C-terminal domain, these covalent bonds fix two long loops on the surface of the enzyme that cover the substrate access channel to the active site. These features provide an explanation for the dual activities of this enzyme.

Amino Acid Sequence↗

Cluster control of a distributed-parameter planar structure--middle authority control.

This paper proposes a generalized active vibration control approach, "cluster control," that explicitly targets sets of structural modes with some common property. The approach falls into a category of MAC (middle authority control) between conventionally used LAC (low authority control) and HAC (high authority control), possessing the benefit of stability and control law simplicity analogous to LAC, while providing high control performance as well as some flexibility of control gain assignment similar to HAC. The structure of a cluster control system is outlined, showing that it is possible to control a target cluster without affecting the other clusters. A design procedure for the cluster control system is then proposed. Experimental results are also presented.

Journal Article↗

Selective surface modification technique for improvement of chromatographic separation selectivity for sugar derivatives.

Highly cross-linked macroporous polymers were prepared utilizing ethylene dimethacrylate as a cross-linking agent, in the presence or absence of methyl-alpha-D-glucoside as a kind of template molecule with methacrylic acid as a functional monomer. After the preparation of the polymers, we applied a high temperature to the cross-linked polymers to study the changes of adsorption properties of the polymers for sugar derivatives including the template molecule utilized. Interestingly, the heat treatment up to 250 degrees C afforded improvement of relative adsorption affinity for several sugar derivatives including the template molecule, while heat treatment up to 150 degrees C did not afford those improvements. The detailed studies including polymers prepared using acrylic acid as a functional monomer instead of methacrylic acid prove that temperatures higher than the Tg temperature of the polymer derived from a functional monomer such as methacrylic acid and higher than the melting point (mp) of the sugar template are necessary to afford the observed improvement of relative affinity based on the surface modification effects through the heat treatment to cross-linked polymers.

Carbohydrates↗