2006 Fiscal Year Final Research Report Summary
The Single Molecular Design and Systemic Assembly of Protein Function
| Project/Area Number |
14002008
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| Research Category |
Grant-in-Aid for Specially Promoted Research
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| Allocation Type | Single-year Grants |
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| Review Section |
Biological Sciences
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| Research Institution | Waseda University |
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Principal Investigator |
ISHIWATA Shinichi Waseda University, Faculty of Science and Engineering, Professor, 理工学術院, 教授 (10130866)
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| Co-Investigator(Kenkyū-buntansha) |
FUNATSU Takashi The University of Tokyo, Graduate School of Pharmaceutical Sciences, Professor, 薬学研究科(研究院), 教授 (00190124)
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| Project Period (FY) |
2002 – 2006
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| Keywords | Biomolecular motor / Bionanogauge / Optical Tweezers / SPOC / Microthermometer / Photon Counting Histogram / Actin / Mitotic Spindle |
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| Research Abstract |
Biological functions are inherently hierarchical. We planned to investigate the underlying mechanism of biological functions from the single-molecular level to the level of supramolecular assemblies, focusing on the mechanism characteristic of each level of hierarchy. Our study was focused primarily on a biological motility system starting from interactions between single-molecular motors and track polymers, to whole ensembles of such interactions (nanomuscle, myofibrils and fibers), and on to single cells and whole tissues (the heart). We also aimed to develop a technique that will allow the use of the cytoskeleton as a "bionanogauge". The main results of this project are summarized as follows: 1) Microscopic measurements of unbinding force of actin-myosin V or VI complex under various concentrations of ADP showed that the apparent binding affinity of ADP depends on the loading direction. Based on a simple model analysis, we could determine the load dependence of the attachment and detachment rate constants of ADP to actomyosin complex, which helped understanding of the molecular mechanism of myosin motility. 2) We have demonstrated that the fluorescence intensity of rhodamine-labeled single actin filaments changes depending on the applied load, implying that the local structure of actin is distorted. 3) We presented evidence for that the changes of filament lattice are responsible for the mechanism of SPOC of myofibrils. 4) We have succeeded in identifying the amino acids of tubulin that are responsible for the strong binding of kinesin using genetically engineered tubulin molecules. 5) Using a micro-thermometer, we could find that the application of ionomycin induced the temperature increase by 1℃ in single HeLa cells, which is attributable to the Ca^<2+> uptake. 6) Using the Photon Counting Histogram technique, we showed that actin filaments consist of not only helical polymers but also linear polymers as theoretically predicted by F. Oosawa and his associates.
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[Journal Article] Identification of a strong binding site for kinesin on the microtubule using mutant analysis of tubulin2006
Author(s)
Uchimura, S., Oguchi, Y., Katsuki, M., Usui, T., Osada, H., Nikawa, J., Ishiwata, S., Muto, E.
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Journal Title
EMBO J. 25
Pages: 5932-5941
Description
「研究成果報告書概要(欧文)」より
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[Journal Article] Synthesis of the isotope-labeled derivatization reagent for carboxylic acids, 7-(N, N-dimethylaminosulfony1)-4-(aminoethyl)piperazino-2,1,3-benzoxadiazole (d_6) (DBD-PZ-NH_2 (D)), and its application to the quantification and the determination of relative amount of fatty acids in rat plasma samples by high-performance liquid chromatography/mass spectrometry2006
Author(s)
Tsukamoto Y, T.Santa, H.Yoshida, H.Miyano, T.Fukushima, K.Hirayama, K.Imai, T.Funatsu.
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Journal Title
Biomedical Chromatography 20
Pages: 358-364
Description
「研究成果報告書概要(欧文)」より
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[Journal Article] Suppression of thiol exchange reaction in the determination of reduced-form thiols by high-performance liquid chromatography with fluorescence detection after derivatization with fluorogenic benzofurazan reagent, 7-fluoro-2,1,3-benzoxadiazole-4-sulfonate and 4-aminosulfony1-7-fluoro-2,1,3- benzoxadiazole2006
Author(s)
Santa, T., C.Aoyama, T.Fukushima, K.Imai, T.Funatsu.
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Journal Title
Biomed Chromatogr. 20
Pages: 656-661
Description
「研究成果報告書概要(欧文)」より
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[Journal Article] Higashinakagawa. Cellular responses of the ciliate, Tetrahymena thermophila, to far infrared irradiation2006
Author(s)
Shiurba, R., T.Hirabayashi, M.Masuda, A.Kawamura, Y.Komoike, W.Klitz, K.Kinowaki, T.Funatsu, S.Kondo, S.Kiyokawa, T.Sugai, K.Kawamura, H.Namiki, T.
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Journal Title
Photochem. Photobiol. Sci. 5
Pages: 799-807
Description
「研究成果報告書概要(欧文)」より
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