2013 Fiscal Year Final Research Report
Development of experimental techniques for the investigation of transient molecular protein complexes
| Project Area | Integrative understanding of biological processes mediated by transient macromolecular complexes; New technology for visualizing physiologically metastable states. |
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| Project/Area Number |
21121003
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| Research Category |
Grant-in-Aid for Scientific Research on Innovative Areas (Research in a proposed research area)
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| Allocation Type | Single-year Grants |
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| Review Section |
Complex systems
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| Research Institution | Kyushu University |
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Principal Investigator |
KOHDA Daisuke 九州大学, 生体防御医学研究所, 教授 (80186618)
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| Co-Investigator(Renkei-kenkyūsha) |
SHIMADA Atsushi 九州大学, 生体防御医学研究所, 准教授 (70391977)
MAYANAGI Kouta 九州大学, 生体防御医学研究所, 助教 (50418571)
SAITOH Takashi 九州大学, 生体防御医学研究所, 助教 (00432914)
IGURA Mayumi 九州大学, 生体防御医学研究所, 学術研究院 (60529965)
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| Project Period (FY) |
2009-07-23 – 2014-03-31
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| Keywords | タンパク質 / 分子認識 / プレ配列 / Tom20 / オリゴ糖転移酵素 / Ｘ線結晶解析 / ＮＭＲ解析 / 複合体 |
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| Research Abstract |
Equilibrium shifting from the unbound states to the bound states is useful to obtain structural and dynamic information of transient protein complexes at atomic resolutions, in particular, the complexes with large amplitude motions inside. We developed two techniques, molecular tethering (via inter-molecular disulfide bonds) and molecular stiffening (via intra-molecular disulfide bonds), for this purpose. We also proposed intentional creation of space in protein crystal lattice, and place a target region in the space created. The key technique is the rigid connection to fix the relative orientation of the protein of interest and a tag protein. These new techniques were applied to analyze the dynamics of a mitochondrial presequence peptide as bound to the mitochondrial receptor Tom20, and the stabilization of oligosaccharyltransferase-peptide complexes for crystallization.
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Research Products
(23 results)
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[Journal Article] Structural elucidation of an asparagine-linked oligosaccharide from the hyperthermophilic archaeon, Pyrococcus furiosus2014
Author(s)
Fujinami, D., Matsumoto, M., Noguchi, T., Sonomoto, K., Kohda, D.
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Journal Title
Carbohydr Res
Volume: 387C
Pages: 30-36
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[Journal Article] Energetics of the presequence-binding poses in mitochondrial protein import through Tom202013
Author(s)
Komuro, Y., Miyashita, N., Mori, T., Muneyuki, E., Saitoh, T., Kohda, D., Sugita, Y.
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Journal Title
J Phys Chem B
Volume: 117
Pages: 2864-2871
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[Journal Article] Crystallographic and NMR evidence for flexibility in oligosaccharyltransferases and its catalytic significance2013
Author(s)
Nyirenda, J., Matsumoto, S., Saitoh, T., Maita, N., Noda, N. N., Inagaki, F., Kohda, D.
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Journal Title
Structure
Volume: 21
Pages: 32-41
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[Journal Article] Crystal structures of an archaeal oligosaccharyltransferase provide insights into the catalytic cycle of N-linked protein glycosylation2013
Author(s)
Matsumoto, S., Shimada, A., Nyirenda, J., Igura, M., Kawano, Y., Kohda, D.
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Journal Title
Proc Natl Acad Sci U S A
Volume: 110
Pages: 17868-17873
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[Journal Article] Crystal structure of the C-terminal globular domain of oligosaccharyltransferase from Archaeoglobus fulgidus at 1.75 A resolution2012
Author(s)
Matsumoto, S., Igura, M., Nyirenda, J., Matsumoto, M., Yuzawa, S., Noda, N., Inagaki, F., Kohda, D.
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Journal Title
Biochemistry
Volume: 51
Pages: 4157-4166
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[Journal Article] Crystallographic snapshots of Tom20-mitochondrial presequence interactions with disulfide-stabilized peptides2011
Author(s)
Saitoh, T., Igura, M., Miyazaki, Y., Ose, T., Maita, N., Kohda, D.
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Journal Title
Biochemistry
Volume: 50
Pages: 5487-5496
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