24 results on '"Komoto N"'
Search Results
2. Viscous stress–strain behaviour of clay under unloaded conditions
- Author
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Li, J, primary, Nishi, T, additional, Komoto, N, additional, and Tatsuoka, F, additional
- Published
- 2003
- Full Text
- View/download PDF
3. Microstructure control for developing Fe–Pd ferromagnetic shape memory alloys
- Author
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Yasuda, H.Y., Komoto, N., Ueda, M., and Umakoshi, Y.
- Published
- 2002
- Full Text
- View/download PDF
4. Gene introduction into silkworm embryos with a pseudotyped retroviral vector
- Author
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Komoto, N., Thibert, C., Boulo, V., J.C., Burns, Couble, P., and Laviron, Nathalie
- Subjects
[SDV.BC] Life Sciences [q-bio]/Cellular Biology - Published
- 2000
5. ChemInform Abstract: CYCLOADDITION OF IMIDAZO(2,1‐B)THIAZOLES AND THIAZOLO(3,2‐A)BENZIMIDAZOLE WITH DIMETHYL ACETYLENEDICARBOXYLATE
- Author
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ABE, N., primary, NISHIWAKI, T., additional, and KOMOTO, N., additional
- Published
- 1980
- Full Text
- View/download PDF
6. ChemInform Abstract: STUDIES ON HETEROPENTALENES. II. CYCLOADDITION OF IMIDAZO(2,1‐B)THIAZOLES, THIAZOLO(3,2‐A)BENZIMIDAZOLE, AND IMIDAZO(2,1‐B)BENZOTHIAZOLE WITH A REACTIVE ACETYLENIC ESTER
- Author
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ABE, N., primary, NISHIWAKI, T., additional, and KOMOTO, N., additional
- Published
- 1981
- Full Text
- View/download PDF
7. Cluster Analysis of Subjective Shoulder Stiffness and Muscle Hardness: Associations with Central Sensitization-Related Symptoms.
- Author
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Komoto N, Sakebayashi H, Imagawa N, Mizuno Y, Nakata I, Shigetoh H, Kodama T, and Miyazaki J
- Subjects
- Humans, Hardness, Cross-Sectional Studies, Muscle, Skeletal, Shoulder Pain etiology, Cluster Analysis, Shoulder, Central Nervous System Sensitization
- Abstract
Background and Objectives : Understanding the relationships between subjective shoulder stiffness, muscle hardness, and various factors is crucial. Our cross-sectional study identified subgroups of shoulder stiffness based on symptoms and muscle hardness and investigated associated factors. Materials and Methods : measures included subjective stiffness, pain, muscle hardness, and factors like physical and psychological conditions, pressure pain threshold, postural alignment, heart rate variability, and electroencephalography in 40 healthy young individuals. Results : Three clusters were identified: Cluster 1 with high stiffness, pain, and muscle hardness; Cluster 2 with low stiffness and pain but high muscle hardness; and Cluster 3 with low levels of all factors. Cluster 1 had significantly higher central sensitization-related symptoms (CSS) scores than Cluster 2. Subjective stiffness is positively correlated with psychological factors. Conclusions : our results suggest that CSS impacts subjective symptom severity among individuals with similar shoulder muscle hardness.
- Published
- 2023
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- View/download PDF
8. The Impact of Stretching Intensities on Neural and Autonomic Responses: Implications for Relaxation.
- Author
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Imagawa N, Mizuno Y, Nakata I, Komoto N, Sakebayashi H, Shigetoh H, Kodama T, and Miyazaki J
- Subjects
- Humans, Young Adult, Autonomic Nervous System physiology, Electroencephalography, Exercise, Sympathetic Nervous System physiology, Cross-Over Studies, Hamstring Muscles physiology, Muscle Stretching Exercises
- Abstract
Stretching is an effective exercise for increasing body flexibility and pain relief. This study investigates the relationship between stretching intensity and relaxation effects, focusing on brainwaves and autonomic nervous system (ANS) activity. We used a crossover design with low- and high-intensity conditions to elucidate the impact of varying stretching intensities on neural activity associated with relaxation in 19 healthy young adults. Participants completed mood questionnaires. Electroencephalography (EEG) and plethysmography measurements were also obtained before, during, and after stretching sessions. The hamstring muscle was targeted for stretching, with intensity conditions based on the Point of Discomfort. Data analysis included wavelet analysis for EEG, plethysmography data, and repeated-measures ANOVA to differentiate mood, ANS activity, and brain activity related to stretching intensity. Results demonstrated no significant differences between ANS and brain activity based on stretching intensity. However, sympathetic nervous activity showed higher activity during the rest phases than in the stretch phases. Regarding brain activity, alpha and beta waves showed higher activity during the rest phases than in the stretch phases. A negative correlation between alpha waves and sympathetic nervous activities was observed in high-intensity conditions. However, a positive correlation between beta waves and parasympathetic nervous activities was found in low-intensity conditions. Our findings suggest that stretching can induce interactions between the ANS and brain activity.
- Published
- 2023
- Full Text
- View/download PDF
9. Large-Scale Molecular Dynamics Simulation for Ground and Excited States Based on Divide-and-Conquer Long-Range Corrected Density-Functional Tight-Binding Method.
- Author
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Komoto N, Yoshikawa T, Nishimura Y, and Nakai H
- Abstract
In this study, divide-and-conquer (DC) based density-functional tight-binding (DFTB) and time-dependent density-functional tight-binding (TD-DFTB) methods were developed using long-range correction (LC), which resolved the underestimation of energy gaps between the highest occupied molecular orbital and lowest unoccupied molecular orbital. We implemented the LC term by the entrywise product for the effective utilization of the math kernel library. Test calculations of formaldehyde in explicit water molecules demonstrate the efficiency of the developed method. Furthermore, the DC-TD-LCDFTB method was applied to 2,2'-bipyridine-3,3'-diol (BP(OH)
2 ), which exhibits excited-state intramolecular proton transfer in polar solvents.- Published
- 2020
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10. GPU-Accelerated Large-Scale Excited-State Simulation Based on Divide-and-Conquer Time-Dependent Density-Functional Tight-Binding.
- Author
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Yoshikawa T, Komoto N, Nishimura Y, and Nakai H
- Abstract
The present study implemented the divide-and-conquer time-dependent density-functional tight-binding (DC-TDDFTB) code on a graphical processing unit (GPU). The DC method, which is a linear-scaling scheme, divides a total system into several fragments. By separately solving local equations in individual fragments, the DC method could reduce slow central processing unit (CPU)-GPU memory access, as well as computational cost, and avoid shortfalls of GPU memory. Numerical applications confirmed that the present code on GPU significantly accelerated the TDDFTB calculations, while maintaining accuracy. Furthermore, the DC-TDDFTB simulation of 2-acetylindan-1,3-dione displays excited-state intramolecular proton transfer and provides reasonable absorption and fluorescence energies with the corresponding experimental values. © 2019 Wiley Periodicals, Inc., (© 2019 Wiley Periodicals, Inc.)
- Published
- 2019
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11. Development of Large-Scale Excited-State Calculations Based on the Divide-and-Conquer Time-Dependent Density Functional Tight-Binding Method.
- Author
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Komoto N, Yoshikawa T, Ono J, Nishimura Y, and Nakai H
- Abstract
In this study, the divide-and-conquer (DC) method was extended to time-dependent density functional tight-binding (TDDFTB) theory to enable excited-state calculations of large systems and is denoted by DC-TDDFTB. The efficient diagonalization algorithms of TDDFTB and DC-TDDFTB methods were implemented into our in-house program. Test calculations of polyethylene aldehyde and p-coumaric acid, a pigment in photoactive yellow protein, in water demonstrate the high accuracy and efficiency of the developed DC-TDDFTB method. Furthermore, the (TD)DFTB metadynamics simulations of acridinium in the ground and excited states give reasonable p K
a values compared with the corresponding experimental values.- Published
- 2019
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12. Acyl flavonoids, biflavones, and flavonoids from Cephalotaxus harringtonia var. nana.
- Author
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Komoto N, Nakane T, Matsumoto S, Hashimoto S, Shirota O, Sekita S, and Kuroyanagi M
- Subjects
- Chromatography, High Pressure Liquid, Humans, Magnetic Resonance Spectroscopy, Molecular Structure, Plant Leaves chemistry, Biological Products chemistry, Flavonoids chemistry, Harringtonines chemistry
- Abstract
A methanol extract of the leaves of Cephalotaxus harringtonia var. nana and its ethyl acetate (EtOAc)-soluble fraction demonstrated strong antitumor activity against A549 and HT-29 cell lines. The EtOAc-soluble fraction was purified by column chromatography and high-performance liquid chromatography (HPLC) using a reverse-phase column to yield three novel acyl flavonoids and a biflavonoid, along with 15 other known compounds that included flavonoids, biflavonoids, and other phenolics. The structures of the new compounds were elucidated using spectral data from HR-MS and NMR, including two-dimensional NMR studies, as (2R,3R)-3-O-eicosanoyltaxifolin (1), (2R,3R)-3-O-docosanoyltaxifolin (2), (2R,3R)-3-O-tetracosanoyltaxifolin (3), and 6-methyl-4',7,7″-tri-O-methylamentoflavone (4). The isolated compounds, including the known compounds, were tested for possible antitumor activity; some of the biflavones were found to be active. The potent antitumor activity of the extract was attributed to Cephalotaxus alkaloids, such as homoharringtonine (20).
- Published
- 2015
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13. Serotonergic modulation of neuronal activity in the nucleus accumbens following repeated methamphetamine administration in rats.
- Author
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Ishihara K, Takahashi N, Komoto N, Yoshikawa C, Fukumoto S, Ide S, Kimura T, and Ozawa K
- Subjects
- Animals, Enzyme Activators pharmacology, In Vitro Techniques, Male, Methamphetamine administration & dosage, Neural Inhibition drug effects, Nucleus Accumbens physiology, Rats, Rats, Wistar, Second Messenger Systems physiology, Serotonin physiology, Serotonin Receptor Agonists pharmacology, Time Factors, Adenylyl Cyclases physiology, Electrophysiological Phenomena drug effects, Methamphetamine pharmacology, Nucleus Accumbens drug effects, Receptors, Serotonin physiology, Serotonin pharmacology, Serotonin Antagonists, Signal Transduction drug effects, Signal Transduction physiology
- Abstract
Electrophysiological studies were performed to determine whether serotonergic modulation in the nucleus accumbens (NAcc) was affected after repeated methamphetamine (MAP) administration. NAcc slices (400 μm) from Wistar rats administered MAP (5 mg/kg) or saline once daily for 5 days were prepared 1, 5, or 10 days after the final injection. Population spikes (PS) induced by local stimulation of NAcc were recorded. PS inhibition by serotonin was significantly attenuated in the MAP group at 5 days but did not differ at 1 or 10 days. We next analyzed the effects of serotonin receptor subtype (5-HT1A,2,3,4,6,7)-selective agonists of PS. Differences between saline and MAP groups in 5-HT1A,2,3,4,6 receptor agonist-induced changes of PS were small or not significant. Interestingly, 5-HT7 receptor agonists significantly enhanced PS in the MAP group. Changes in the secondary messenger system related to 5-HT7 receptors were also investigated. Adenylate cyclase activator-induced PS enhancements were significantly larger in the MAP group. However, dibutyryl-cAMP-induced PS enhancement was not significantly different. In conclusion, 5-HT-induced inhibition of PS in NAcc was attenuated 5 days after repeated MAP treatment: the change in the effect of 5-HT was probably due to enhancement of the excitatory modulation via the 5-HT7 receptor with adenylate cyclase signal transduction systems.
- Published
- 2013
- Full Text
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14. Murine metabolism and absorption of lancemaside A, an active compound in the roots of Codonopsis lanceolata.
- Author
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Komoto N, Ichikawa M, Ohta S, Nakano D, Nishihama T, Ushijima M, Kodera Y, Hayama M, Shirota O, Sekita S, and Kuroyanagi M
- Subjects
- Administration, Oral, Animals, Cecum chemistry, Cecum metabolism, Chromatography, Liquid, Esters chemistry, Esters metabolism, Male, Mass Spectrometry, Mice, Molecular Structure, Oleanolic Acid analogs & derivatives, Oleanolic Acid chemistry, Oleanolic Acid metabolism, Saponins blood, Saponins chemistry, Saponins pharmacokinetics, Codonopsis chemistry, Plant Roots chemistry, Saponins metabolism
- Abstract
Lancemaside A, a triterpenoid saponin isolated from the roots of Codonopsis lanceolata, has been reported to ameliorate the reduction of blood testosterone levels induced by immobilization stress in mice. In the present study, we investigated the metabolism and absorption of lancemaside A in mice. After oral administration of lancemaside A at 100 mg/kg body weight, the unmetabolized compound appeared rapidly in plasma (t (max) = 0.5 h). Lancemaside A has a low bioavailability (1.1%) because of its metabolism by intestinal bacteria and its poor absorption in the gastrointestinal tract. Furthermore, we identified four metabolites from the cecum of mice after oral administration of lancemaside A: codonolaside II, echinocystic acid, echinocystic acid 28-O-beta-D: -xylopyranosyl-(1 --> 4)-alpha-L: -rhamnopyranosyl-(1 --> 2)-alpha-L: -arabinopyranosyl ester, and echinocystic acid 28-O-alpha-L: -rhamnopyranosyl-(1 --> 2)-alpha-L: -arabinopyranosyl ester. Among these metabolites, codonolaside II and echinocystic acid were detected in plasma, and their t (max) values were 4 and 8 h, respectively. These findings should be helpful for understanding the mechanism of the biological effect of lancemaside A.
- Published
- 2010
- Full Text
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15. Proptosis in dysthyroid ophthalmopathy: a case series of 10,931 Japanese cases.
- Author
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Kozaki A, Inoue R, Komoto N, Maeda T, Inoue Y, Inoue T, and Ayaki M
- Subjects
- Adolescent, Adult, Aged, Aged, 80 and over, Asian People, Child, Child, Preschool, Diagnostic Techniques, Ophthalmological, Exophthalmos diagnosis, Eyelid Diseases diagnosis, Female, Graves Ophthalmopathy diagnosis, Humans, Hypertrophy, Magnetic Resonance Imaging, Male, Middle Aged, Oculomotor Muscles pathology, Tomography, X-Ray Computed, Young Adult, Exophthalmos complications, Graves Ophthalmopathy complications
- Abstract
Purpose: This observational case series investigated the occurrence and distribution of proptosis in Japanese patients with dysthyroid ophthalmopathy (DO). The relationship among lid retraction, lid swelling, and enlargement of extraocular muscles was also analyzed., Methods: From 1993 to 2002, 10 931 patients (2240 men and 8691 women, average age; 39.9 ± 14.0 years) with abnormal thyroid function were examined for proptosis. Proptosis was measured using a Hertel exophthalmometer, and lid swelling and retraction were classified. Orbital magnetic resonance imaging or computed tomography was used to quantify enlargement of the extraocular muscles., Results: Clinically significant proptosis (≥15 mm) was present in 74.2% patients and was classified as mild (15-17 mm, 39.1%), moderate (18-20 mm, 25.9%), or severe (>20 mm, 9.2%). The average exophthalmometer reading was 17.2 ± 3.2 mm, with proptosis less pronounced with age (p < 0.001) and more pronounced in men than women (18.2 ± 3.4 mm vs. 17.0 ± 3.1 mm; p < 0.001). Lid retraction was present in 57.7% of patients, lid swelling in 46.9%, and muscle enlargement in 40.8% of patients. Multiple regression analysis revealed proptosis was strongly correlated with a young age and the male gender, followed by extraocular muscle enlargement and lid retraction and swelling (p < 0.001)., Conclusions: This study assessed the measurable features of proptosis in Japanese patients with DO and contributes to the understanding by correlating symptoms and signs of DO.
- Published
- 2010
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16. Simultaneous determination of seven saponins in the roots of Codonopsis lanceolata by liquid chromatography-mass spectrometry.
- Author
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Ichikawa M, Ohta S, Komoto N, Ushijima M, Kodera Y, Hayama M, Shirota O, Sekita S, and Kuroyanagi M
- Subjects
- Molecular Structure, Reproducibility of Results, Saponins chemistry, Triterpenes analysis, Triterpenes chemistry, Chromatography, High Pressure Liquid methods, Codonopsis chemistry, Plant Roots chemistry, Saponins analysis, Spectrometry, Mass, Electrospray Ionization methods
- Abstract
We developed a rapid and simple analytical method for the simultaneous determination of seven 3,28-bidesmosidic triterpenoid saponins in the roots of Codonopsis lanceolata. The saponins are lancemaside A, lancemaside B, lancemaside C, lancemaside E, lancemaside G, foetidissimoside A, and aster saponin Hb. Root samples were extracted with 50% methanol and prepared for analysis. Saponins were detected by reversed-phase high-performance liquid chromatography with electrospray ionization mass spectrometry, and ginsenoside Rb(1) was used as an internal standard. The overall recoveries of all saponins were 92-116%, and the relative standard deviation values of intra- and inter-day precision were lower than 3.7 and 7.7%, respectively. Eight root samples collected from Korea and Japan were analyzed using the developed method. Lancemaside A was the most abundant saponin in the root samples from Korea, ranging from 2.65 to 3.64 mg/g dry root. However, the maximum content of lancemaside A among Japanese samples was 0.101 mg/g dry root.
- Published
- 2009
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17. Rapid identification of triterpenoid saponins in the roots of Codonopsis lanceolata by liquid chromatography-mass spectrometry.
- Author
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Ichikawa M, Ohta S, Komoto N, Ushijima M, Kodera Y, Hayama M, Shirota O, Sekita S, and Kuroyanagi M
- Subjects
- Chromatography, Liquid methods, Mass Spectrometry methods, Plant Roots, Saponins chemistry, Triterpenes chemistry, Codonopsis chemistry, Saponins isolation & purification, Triterpenes isolation & purification
- Abstract
Liquid chromatography coupled with sequential mass spectrometry (LC-MS(n)) has been used to identify 3,28-bidesmosidic triterpenoid saponins, lancemaside A (1), foetidissimoside A (2), aster saponin Hb (3), lancemaside E (4), lancemaside B (5), lancemaside F (6), lancemaside G (7), lancemaside C (8), and lancemaside D (9) in the roots of Codonopsis lanceolata. Structural information about both the aglycone and the sugar moiety at the C-3 position of saponins was obtained in the negative-ion mode. On the other hand, positive-ion spectra mainly provide structural information about the sugar chains of saponins, especially the oligosaccharide moiety at the C-28 position. During subsequent fragmentation of the product ions derived from the oligosaccharide moiety at the C-28 position, fragments produced by sequential loss of a monosaccharide unit were observed. Furthermore, the structural features of two unknown saponins in the roots of C. lanceolata were assigned on the basis of the fragmentation patterns of the known saponins. These studies demonstrate that LC-MS(n) analysis has great potential for the identification and characterization of triterpenoid saponins in plant extracts.
- Published
- 2008
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18. Preparative separation of the saponin lancemaside a from Codonopsis lanceolata by centrifugal partition chromatography.
- Author
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Shirota O, Nagamatsu K, Sekita S, Komoto N, Kuroyanagi M, Ichikawa M, Ohta S, and Ushijima M
- Subjects
- Magnetic Resonance Spectroscopy, Plant Extracts chemistry, Spectrophotometry, Ultraviolet, Chromatography, High Pressure Liquid methods, Codonopsis chemistry, Saponins isolation & purification
- Abstract
Introduction: Lancemaside A is a saponin that inhibits decreases in blood testosterone level and thus prevents or ameliorates symptoms associated with male climacteric disorder. Our initial attempt to preparative isolation of lancemaside A from the saponin fraction of Codonopsis lanceolata roots by a preparative HPLC did not give a clear result., Objective: To develop a simple and efficient method for the preparative isolation of lancemaside A from the hot water extract of C. lanceolata roots using centrifugal partition chromatography (CPC)., Methodology: The saponin fraction obtained from the hot water extract of C. lanceolata roots was used as the sample for preparative-scale separation of lancemasides by CPC using n-hexane:n-butanol:methanol:0.1% aqueous formic acid (3:4:1:6, v/v) as the two-phase solvent system. The upper phase (organic phase) of the two-phase solvent system was used as the mobile phase, and 0.5 g of saponin fraction was applied for separation by CPC. Each fraction that was separated by CPC was analysed by HPLC, and the fractions containing each of the separated compounds were pooled together, and then were purified by simple preparative HPLC., Results: The demonstrated separation sequence, hot water extraction, DIAION HP-20 column chromatography, CPC and preparative HPLC, yielded lancemaside A, foetidissimoside A and astersaponin Hb in their pure forms., Conclusion: The simple and efficient method for the preparative isolation of lancemaside A along with two other saponins, foetidissimoside A and astersaponin Hb, from the saponin fraction of C. lanceolata was established using CPC.
- Published
- 2008
- Full Text
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19. Histidine-alpha143 assists 1,2-hydroxyl group migration and protects radical intermediates in coenzyme B12-dependent diol dehydratase.
- Author
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Kinoshita K, Kawata M, Ogura K, Yamasaki A, Watanabe T, Komoto N, Hieda N, Yamanishi M, Tobimatsu T, and Toraya T
- Subjects
- Crystallography, X-Ray, Electron Spin Resonance Spectroscopy, Histidine chemistry, Histidine genetics, Hydrogen Bonding, Hydrogen-Ion Concentration, Kinetics, Klebsiella oxytoca enzymology, Klebsiella oxytoca genetics, Klebsiella oxytoca metabolism, Models, Biological, Molecular Structure, Mutagenesis, Site-Directed, Mutation, Propanediol Dehydratase chemistry, Propanediol Dehydratase genetics, Structure-Activity Relationship, Substrate Specificity, Cobamides metabolism, Histidine metabolism, Propanediol Dehydratase metabolism
- Abstract
Diol dehydratase of Klebsiella oxytoca contains an essential histidine residue. Its X-ray structure revealed that the migrating hydroxyl group on C2 of substrate is hydrogen-bonded to Hisalpha143. Mutant enzymes in which Hisalpha143 was mutated to another amino acid residue were expressed in Escherichia coli, purified, and examined for enzymatic activity. The Halpha143Q mutant was 34% as active as the wild-type enzyme. Halpha143A and Halpha143L showed only a trace of activity. Kinetic analyses indicated that the hydrogen bonding interaction between the hydroxyl group on C2 of substrate and the side chain of residue alpha143 is important not only for catalysis but also for protecting radical intermediates. Halpha143E and Halpha143K that did not exist as (alphabetagamma) 2 complexes were inactive. The deuterium kinetic isotope effect on the overall reaction suggested that a hydrogen abstraction step is fully rate-determining for the wild type and Halpha143Q and partially rate-determining for Halpha143A. The preference for substrate enantiomers was reversed by the Halpha143Q mutation in both substrate binding and catalysis. Upon the inactivation of the Halpha143A holoenzyme by 1,2-propanediol, cob(II)alamin without an organic radical coupling partner accumulated, 5'-deoxyadenosine was quantitatively formed from the coenzyme adenosyl group, and the apoenzyme itself was not damaged. This inactivation was thus concluded to be a mechanism-based inactivation. The holoenzyme of Halpha143Q underwent irreversible inactivation by O 2 in the absence of substrate at a much lower rate than the wild type.
- Published
- 2008
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20. Triterpene glycosides from the roots of Codonopsis lanceolata.
- Author
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Ushijima M, Komoto N, Sugizono Y, Mizuno I, Sumihiro M, Ichikawa M, Hayama M, Kawahara N, Nakane T, Shirota O, Sekita S, and Kuroyanagi M
- Subjects
- Carbohydrate Conformation, Glycosides isolation & purification, Hydrolysis, Korea, Magnetic Resonance Spectroscopy, Molecular Conformation, Plant Roots chemistry, Saponins chemistry, Saponins isolation & purification, Spectrometry, Mass, Electrospray Ionization, Triterpenes isolation & purification, Codonopsis chemistry, Glycosides chemistry, Triterpenes chemistry
- Abstract
In the course of the development of new designer foods using the roots of Codonopsis lanceolata, we found that hot-water extracts of C. lanceolata recovered decreased testosterone levels in the blood and accelerated the restoration of reproductive dysfunction induced by hyperthermic treatment in male mice. Thus we studied the constituents of the polar fraction of the roots of C. lanceolata and identified six new triterpene saponins, lancemasides B (2), C (3), D (4), E (5), F (6), and G (7), along with the known saponin lancemasaide A (1) and phenylpropanoid glycosides 8-10. The structures of the new compounds 2-7 were determined by means of spectral data including 2D-NMR studies and chemical reactions to be oleanan-type bisdesmoside with sugars at C-3 and C-28. Compounds 2-6 have echinocystic acid as an aglycone, and compound 7 has asterogenic acid as an aglycone. Identification of the sugars and determination of their D,L-chiralities were carried out by application of the exciton chirality method to the per-O-p-bromobenzoylmethyl sugar derived from saponins.
- Published
- 2008
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21. Construction and characterization of hybrid dehydratases between adenosylcobalamin-dependent diol and glycerol dehydratases.
- Author
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Sakai T, Yamasaki A, Toyofuku S, Nishiki T, Yunoki M, Komoto N, Tobimatsu T, and Toraya T
- Subjects
- Amino Acid Sequence, Bacterial Proteins chemistry, Bacterial Proteins genetics, Cobamides chemistry, Cobamides genetics, Electrophoresis, Polyacrylamide Gel methods, Enzyme Activation genetics, Escherichia coli genetics, Gene Expression, Glycerol chemistry, Hydro-Lyases chemistry, Hydro-Lyases genetics, Klebsiella oxytoca chemistry, Klebsiella oxytoca enzymology, Klebsiella pneumoniae chemistry, Klebsiella pneumoniae enzymology, Protein Subunits chemistry, Protein Subunits genetics, Protein Subunits metabolism, Recombinant Fusion Proteins chemistry, Recombinant Fusion Proteins genetics, Sequence Analysis, Protein methods, Sequence Homology, Amino Acid, Substrate Specificity, Bacterial Proteins metabolism, Cobamides metabolism, Hydro-Lyases metabolism, Recombinant Fusion Proteins metabolism
- Abstract
Adenosylcobalamin-dependent diol dehydratase and glycerol dehydratase are isofunctional enzymes that catalyze the dehydration of 1,2-diols to the corresponding aldehydes. Although they bear different metabolic roles, both enzymes consist of three different subunits and possess a common (alphabetagamma)2 structure. To elucidate the roles of each subunit, we constructed expression plasmids for the hybrid dehydratases between diol dehydratase of Klebsiella oxytoca and glycerol dehydratase of Klebsiella pneumoniae in all the combinations of subunits by gene engineering techniques. All of the hybrid enzymes were produced in Escherichia coli at high levels, but only two hybrid enzymes consisting of the alpha subunit from glycerol dehydratase and the beta subunits from diol dehydratase showed high activity. The substrate specificity, the susceptibility to inactivation by glycerol, and the monovalent cation specificity of the wild type and hybrid enzymes were primarily determined by the origin of their alpha subunits.
- Published
- 2007
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22. Survey of catalytic residues and essential roles of glutamate-alpha170 and aspartate-alpha335 in coenzyme B12-dependent diol dehydratase.
- Author
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Kawata M, Kinoshita K, Takahashi S, Ogura K, Komoto N, Yamanishi M, Tobimatsu T, and Toraya T
- Subjects
- Aspartic Acid, Bacterial Proteins genetics, Catalytic Domain genetics, Cobamides metabolism, Glutamic Acid, Kinetics, Mutagenesis, Site-Directed, Propanediol Dehydratase genetics, Structure-Activity Relationship, Substrate Specificity, Bacterial Proteins chemistry, Bacterial Proteins metabolism, Klebsiella oxytoca enzymology, Propanediol Dehydratase chemistry, Propanediol Dehydratase metabolism
- Abstract
The importance of each active-site residue in adenosylcobalamin-dependent diol dehydratase of Klebsiella oxytoca was estimated using mutant enzymes in which one of the residues interacting with substrate and/or K(+) was mutated to Ala or another amino acid residue. The Ealpha170A and Dalpha335A mutants were totally inactive, and the Halpha143A mutant showed only a trace of activity, indicating that Glu-alpha170, Asp-alpha335, and His-alpha143 are catalytic residues. The Qalpha141A, Qalpha296A, and Salpha362A mutants showed partial activity. It was suggested from kinetic parameters that Gln-alpha296 is important for substrate binding and Gln-alpha296 and Gln-alpha141 for preventing the enzyme from mechanism-based inactivation. The Ealpha221A, Ealpha170H, and Dalpha335A did not form the (alphabetagamma)(2) complex, suggesting that these mutations indirectly disrupt subunit contacts. Among other Glu-alpha170 and Asp-alpha335 mutants, Ealpha170D and Ealpha170Q were 2.2 +/- 0.3% and 0.02% as active as the wild-type enzyme, respectively, whereas Dalpha335N was totally inactive. Kinetic analysis indicated that the presence and the position of a carboxyl group in the residue alpha170 are essential for catalysis as well as for the continuous progress of catalytic cycles. It was suggested that the roles of Glu-alpha170 and Asp-alpha335 are to participate in the binding of substrate and intermediates and keep them appropriately oriented and to function as a base in the dehydration of the 1,1-diol intermediate. In addition, Glu-alpha170 seems to stabilize the transition state for the hydroxyl group migration from C2 to C1 by accepting the proton of the spectator hydroxyl group on C1.
- Published
- 2006
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23. Germline transformation of the silkworm Bombyx mori L. using a piggyBac transposon-derived vector.
- Author
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Tamura T, Thibert C, Royer C, Kanda T, Abraham E, Kamba M, Komoto N, Thomas JL, Mauchamp B, Chavancy G, Shirk P, Fraser M, Prudhomme JC, and Couble P
- Subjects
- Actins genetics, Aging metabolism, Animals, Animals, Genetically Modified, Base Sequence, Bombyx embryology, Bombyx growth & development, Bombyx metabolism, Crosses, Genetic, Female, Green Fluorescent Proteins, Larva metabolism, Luminescent Proteins analysis, Luminescent Proteins genetics, Male, Moths enzymology, Moths genetics, Mutagenesis, Site-Directed genetics, Promoter Regions, Genetic genetics, Pupa metabolism, Recombination, Genetic genetics, Sequence Analysis, DNA, Terminal Repeat Sequences genetics, Transgenes genetics, Transposases genetics, Transposases metabolism, Bombyx genetics, DNA Transposable Elements genetics, Genetic Vectors genetics, Germ-Line Mutation genetics, Transformation, Genetic genetics
- Abstract
We have developed a system for stable germline transformation in the silkworm Bombyx mori L. using piggyBac, a transposon discovered in the lepidopteran Trichoplusia ni. The transformation constructs consist of the piggyBac inverted terminal repeats flanking a fusion of the B. mori cytoplasmic actin gene BmA3 promoter and the green fluorescent protein (GFP). A nonautonomous helper plasmid encodes the piggyBac transposase. The reporter gene construct was coinjected into preblastoderm eggs of two strains of B. mori. Approximately 2% of the individuals in the G1 broods expressed GFP. DNA analyses of GFP-positive G1 silkworms revealed that multiple independent insertions occurred frequently. The transgene was stably transferred to the next generation through normal Mendelian inheritance. The presence of the inverted terminal repeats of piggyBac and the characteristic TTAA sequence at the borders of all the analyzed inserts confirmed that transformation resulted from precise transposition events. This efficient method of stable gene transfer in a lepidopteran insect opens the way for promising basic research and biotechnological applications.
- Published
- 2000
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24. Gibberellin-antagonizing protein in dwarf peas.
- Author
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Komoto N, Ikegami S, and Tamura S
- Abstract
Seedlings of dwarf pea grown under red light for 9 d were homogenized in 0.1 M phosphate buffer (pH 6.8) and a "water-soluble extract" was obtained by centrifugation, dialysis and lyophilization. The extract contained a proteinaceous substance (or substances) which interfered with the GA3 response of dwarf peas, probably due to complex formation with the hormone.
- Published
- 1973
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