2003 Fiscal Year Final Research Report Summary
A comprehensive study on cytotoxic tRNases
| Project/Area Number |
12306005
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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 |
応用微生物学・応用生物化学
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| Research Institution | The University of Tokyo |
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Principal Investigator |
MASAKI Haruhiko The University of Tokyo, Graduate School of Agricultural and Sciences, Professor (50134515)
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| Co-Investigator(Kenkyū-buntansha) |
HIDAKA Makoto The University of Tokyo, Graduate School of Agricultural and Life Sciences, Associate Professor (50183918)
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| Project Period (FY) |
2000 – 2003
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| Keywords | colicin / tRNA / ribonuclease / tRNase / restriction enzyme / Escherichia coli O157 / molecular mimicry / anticodon |
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| Research Abstract |
We characterized colicins E5 and D as "cytotoxic tRNases", a new type of toxins that specifically cleave tRNA anticodon loops. On the infection in sensitive Escherichia coli, the C-terminal ribonuclease domain of colicin E5, E5-CRD, enters the cell and cleaves anticodons of tRNA^<Tyr>, tRNA^<His>, tRNA^<Asn> and tRNA^<Asp> to kill the cell. Likewise, the C-terminal domain of colicin D, D-CRD, cleaves the 3' terminal of anticodon loops of four is accepting tRNA^<Arg> molecules.
The X-ray crystallographic study and mutation experiments revealed that E5-CRD specifically recognizes the common GU dinucleotide in susceptible tRNA anticodons, by mimicking the structure and function of corresponding codons of mRNA. Thus, E5-CRD can be said as an "RNA restriction enzyme" that cleaves single stranded GU sequences. On the other hand, the inhibitor protein of colicin E5, ImmE5, which originally protects the colicinogenic cells from lethality by the cognate colicin, was shown to bind E5-CRD at the substrate site by mimicking tRNA anticodons. This double molecular mimicry represents a novel process that a pair of proteins has acquired a specific protein-protein interaction by taking over an RNA-RNA interaction through an RNA-protein interaction. We also determined the structures of D-CRD and the D-CRD/ImmD complex, though we failed to visualize interacting structures of D-CRD and the substrate. Since tRNA anticodon loop structures are conserved among a wide range of organisms, E5-CRD and D-CRD function even in eukaryotic cells and are expected to be novel tools to analyze eukaryotic cell mechanisms.
We also screened a collection of E. coli 0157 strains for new type tRNase colicins other than E5 and D. We could not discover such a colicin but found that about 1/4 of the whole O157 collection proved to produce some colicins and that unusually more than 3/4 of those colicins were colicin D.
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Research Products
(26 results)
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[Journal Article] Relation between tRNase activity and the structure of colicin D according to X-ray crystallography.2004
Author(s)
Yajima, S., Nakanishi, K., Takahashi, K., Ogawa, T., Hidaka, M., Kezuka, Y., Nonaka, T., Ohsawa, K., Masaki, H.
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Journal Title
Biochem. Biophys. Res. Com. 966-973
Description
「研究成果報告書概要(欧文)」より
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[Journal Article] In vitro selection of RNA aptamers that bind to colicin E3 and structurally resemble the decoding site of 16S ribosomal RNA.2004
Author(s)
Hirao, I., Harada, Y., Nojima, T., Ogawa, Y., Masaki, M., Yokoyama, S.
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Journal Title
Biochemistry 43
Pages: 3214-3221
Description
「研究成果報告書概要(欧文)」より
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[Presentation] Specific recognition of the substrate tRNAs and inhibitor proteins by colicins E5 and D.2003
Author(s)
Masaki, H., Ogawa, T., Nakanishi, K., Inoue, S., Hidaka, M., Yajima, S.
Organizer
RNA 2003 Kyoto The new frontier of RNA science
Place of Presentation
Kyoto
Year and Date
2003-11-25
Description
「研究成果報告書概要(欧文)」より
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[Presentation] Structures and functions of ribonucleases that cleave specific tRNAs.2003
Author(s)
Masaki, H., Ogawa, T., Inoue, S., Ito, K., Nakanishi, K., Hidaka, M., Yajima, S.
Organizer
76th Annual Meeting of JBS
Place of Presentation
Yokohama
Year and Date
2003-10-17
Description
「研究成果報告書概要(欧文)」より
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