2006 Fiscal Year Final Research Report Summary
Elucidation of structural principal in active state of membrane proteins using fused polycyclic ethers
| Project/Area Number |
16201044
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| Research Category |
Grant-in-Aid for Scientific Research (A)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Living organism molecular science
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| Research Institution | The University of Tokyo |
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Principal Investigator |
TACHIBANA Kazuo The University of Tokyo, Graduate School of Science, Professor (70142081)
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| Co-Investigator(Kenkyū-buntansha) |
FUKUZAWA Seketsu The University of Tokyo, Graduate School of Science, Assistant Professor (40321806)
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| Project Period (FY) |
2004 – 2006
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| Keywords | marine toxin / polycyclic ether / membrane protein / lipid bilayer / Suzuki cross coupling / Gambierol / photo-affinity labeling / helical structure |
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| Research Abstract |
Marine toxins which contain fused polycyclic ethers are promising chemical tools for investigation on membrane protein functions. Our strategy is consisted with following scheme: (i) preparation of simplified fused polycyclic ether library by chemical synthesis; (ii) methodogy development for obtaining structural information of polyether-membrane protein complex model. We succeeded in synthesis of simplified poly ethers based on B-alkyl Suzuki-Miyaura cross coupling and confirmed them recognized a helical structure using CD spectroscopy. For development of membrane protein mimic system we polished up model membrane system using bicelle (DMPC/DHPC: phospholipid aggreregates) system. Characterization of their temperature dependent morphological changes has been done using^<31>P-NMR, DLS, and DSC. Furthermore we developed several protein labeling techniques. A new carbon-carbon bond has been regioselectively introduced into a target position (position 32 or 174) of the Ras protein by two types of organopalladium reactions (Mizoroki-Heck and Sonogashira reactions). An Escherichia coli suppressor tRNA^<Phe> (tRNA^<Phe> cuA) was mis-acylated with 4-iodo-L-phenylalanine by the A294G mutant of E. coli phenylalanyl-tRNA synthetase (G294-PheRS) at a high magnesium ion concentration. The pre-acylated tRNA was added to an E. coli cell-free system to synthesize a Ras protein containing 4-iodo-L-phenylalanine residue at specific target position. The iF-Ras proteins containing 4-iodo-L-phenylalanine residue were subjected to organopalladium reactions with vinylated or propargylated biotin. Site-specific biotinylations of the Ras protein were confirmed by Western blot and LC-MS/MS.
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[Journal Article] Site-specific Functionalization of Proteins by Organopalladium Reactions.2007
Author(s)
Kodama, K., Fukuzawa, S., Nakayama, H., Kigawa. T., Sakamoto, K., Yabuki, T., Matsuda, N., Shirouzu, M., Takio, K., Yokoyama, S., Tachibana, K.
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Journal Title
ChemBioChem 8
Pages: 232-238
Description
「研究成果報告書概要(欧文)」より
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[Journal Article] Thermal analyses of phospholipid mixtures by differential scanning calorimetry and effect of doping with a bolaform amphiphile.2007
Author(s)
Sasaki, R., Sasaki, H., Fukuzawa, S., Kikuchi, J., Hirota, H., Tachibana, K.
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Journal Title
Bull Chem. Soc. Jpn. 80
Pages: 1208-1216
Description
「研究成果報告書概要(欧文)」より
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[Journal Article] Regioselective Carbon-Carbon Bond Formation in Proteins by Palladium-catalysis; New Protein Chemistry by Organometallic Chemistry.2006
Author(s)
Kodama, K., Fukuzawa, S., Nakayama. H., Kigawa, T., Sakamoto, K., Yabuki, T., Matsuda, N., Shirouzu, M., Takio, K., Tachibana, K., Yokoyama, S.
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Journal Title
ChemBioChem 7
Pages: 134-139
Description
「研究成果報告書概要(欧文)」より
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[Journal Article] A New Protein Engineering Approach Combining Chemistry and Biology, Part I; Site-specific Incorporation of 4-iodo-L-phenylalanine in vitro Using Mis-acylated Suppressor tRNA ^<phe>.2006
Author(s)
Kodama, K., Fukuzawa, S., Nakayama, H., Kigawa, T., Sakamoto, K., Yabuki, T., Matsuda, N., Shirouzu, M., Takio, K., Tachibana, K., Yokoyama, S.
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Journal Title
ChemBioChem 7
Pages: 1577-1581
Description
「研究成果報告書概要(欧文)」より
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