| 1.Research Institution | Nagoya University | |
| 2.Research Area | Life Sciences | |
| 3.Research Field | Structure and Functional Control Mechanism of Biomolecules | |
| 4.Term of Project | FY1996〜FY2000 | |
| 5.Project Number | 96L00504 | |
| 6.Title of Project | Structure Recognition and Molecular Mechanism in Bioluminescence |
| Name | Institution,Department | Title of Position |
| Minoru Isobe | Nagoya University, Graduate School of Bioagricultural Sciences | Professor. |
8.Core Members
| Names | Institution,Department | Title of Position |
| Tsukasa Matsuda | Nagoya University, Graduate School of Bioagricultural Sciences | Professor |
| Akira Takai | Nagoya University, Graduate School of Medicine | Assistant Professor |
9.Cooperating Researchers
| Names | Institution,Department | Title of Position |
| Hidenori Kai | Tottori University, Faculty of Agriculture | Professor |
10.Summary of Research Results
|
The photoprotein, symplectin, was obtained from Okinawan squid responsible for bioluminescence, and its 501 amino acids were sequenced through Q-TOF-MS analysis as well as cDNA method. Light production was confirmed by overexpression of this protein through cloning to E. coli. The chromophore that include 100% 13C labeling and several analogs were synthesized and it was recombinated with aposymplectin to provide luminescence, and the molecular mechanism of the luminescent process has been elucidated. Thus, the chromophore binds to protein via covalent bond through a sulfur atom of cystein residue. The recombinant symplectin with a fluorine-containing chromophore afforded evidence of a fluorescent peptide through nano-LC-Q-TOF-MS/MS method to elucidate that the 340 cystein might bind the chromophore. Site specific modification of porteins can be readily monitored by this new methodology, which made protein analysis and modified protein analysis very easy. On the other hand, the analytical method established during this project was applied to other proteinaous system; thus, time interval measurement enzyme is a glycoprotein comprising from 156 amino acid residue with 4 sugar chains through aspargine-22 through N-glycoside linkage. |
11.Key Words
(1)luminous squid、(2)symplectin、(3)cDNA cloning
(4)13C-NMR、(5)chromophore、(6)nano-LC-Q-TOF-MS
(7)EA4、(8)tryptic digestion、(9)glycoprotein
12.References
| Author | Title of Article | |||
| Kuse, M.; Isobe, M. | Synthesis of 13C-Dehydrocoelenterazxine and NMR Studies on Bioluminescence of Symplectoteuthis model | |||
| Journal | Volume | Year | Pages Concerned | |
| Tetrahedron | 56 | 2000 | 2629-2639 | |
| Author | Title of Article | |||
| Takai, A.; Tsuboi, K.; Koyasu, M.; Isobe, M | Effects of modification of the hydrophobic C1-C16 segment of tautomycin on its affinity to type 1 and type 2A protein phosphatases | |||
| Journal | Volume | Year | Pages Concerned | |
| Biochemical Journal | 350 | 2000 | 81-88 | |
| Author | Title of Article | |||
| Liu, Tong-Zhu; Isobe, M. | Synthetic Studies on the HIJK-Ring Fragment of Ciguatoxin | |||
| Journal | Volume | Year | Pages Concerned | |
| Tetrahedron | 56 | 2000 | 5391-5404 | |
| Author | Title of Article | |||
| Suwan, S.; Isobe, M.; Kanokmedhakul, S.; Lourit, N.; Kanokmedhakul, K.; Soytong, K.; Koga, K. | Elucidation of Great Micro-heterogeneity of an Acidic-Neutral Trichotoxin Mixture from Trichoderma harzianum by ESI-QTOF Mass Spectrometry | |||
| Journal | Volume | Year | Pages Concerned | |
| J. Mass Spectrometry | 35 | 2000 | 1438-1451 | |
| Author | Title of Article | |||
| Kuse, M.; Suwan, S.; Koga, K.; Franz, T.; Kanakubo, A.; Isobe, M.; Shimomura, O. | 7, 8-Dihydropterin-6-carboxylic Acid as Light Emitter of Lumonous Millipede, Luminodesmus sequoiae | |||
| Journal | Volume | Year | Pages Concerned | |
| J. Bioorganic Chem. Lett. | 2001 | |||
| Author | Title of Article | |||
| Tani, N.; Kamada, G.; Ochiai, K.; Isobe, M.; Suwan, S.; Kai. H. | Carbohydrate Moiety of Time-Interval Measuring Enzyme Regulates Time Measurement through Its Interaction with Time-Holding Peptide PIN | |||
| Journal | Volume | Year | Pages Concerned | |
| J. Biochem. | 129 | 2001 | 221-227 | |
| Author | Title of Book | ||
| Isobe, M. | Bioscience, Biotechnology, Biochemistry | ||
| Publisher | Year | Pages | |
| the Japan Society for Bioscience, Biotechnology, and Agrochemistry | 2000 | 12 | |