Study on Structure and Function of Vero Toxins (Shiga-like) Toxins Produced by EscherichiaDB coli
| Project/Area Number |
01480174
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| Research Category |
Grant-in-Aid for General Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
細菌学
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| Research Institution | KYOTO UNIVERSITY |
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Principal Investigator |
TAKEDA Yoshifumi Faculty of Medicine, Kyoto University, Professor, 医学部 (30029772)
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| Co-Investigator(Kenkyū-buntansha) |
NISHIBUCHI Mitsuaki Faculty of Medicine, Kyoto University, Associate Professor, 医学部, 助教授 (50189304)
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| Project Period (FY) |
1989 – 1990
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| Project Status |
Completed (Fiscal Year 1990)
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| Budget Amount *help |
¥6,800,000 (Direct Cost: ¥6,800,000)
Fiscal Year 1990: ¥2,400,000 (Direct Cost: ¥2,400,000)
Fiscal Year 1989: ¥4,400,000 (Direct Cost: ¥4,400,000)
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| Keywords | Vero toxin / Enterohemorrhagic E. coli / Inhibition of protein synthesis / RNA N-glycosidase / Site directed mutagenesis / 活性中心 / 腸管出血性大腸菌 / RNA N-グリコシダ-ゼ |
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| Research Abstract |
Enterohemorrhagic Escherichia coli causes infant diarrhea, hemorrhagic colitis and hemolytic uremic syndrome in man. Pathogenesis is considered to be closely related to Vero toxins produced by the organisms. It has been reported that both two types of Vero toxins (VT1 and VT2) show RNA N-glycosidase activity and cleave a glycosidic bond of adenosine at position 4324 from N-terminus of 28S ribosomal RNA of 60S ribosomal subunit of eukaryotic cells. This enzymatic activity results in the inhibition of EF-u dependent tRNA binding and thus the inhibition of protein synthesis. This mode of action to that of ricin.
Comparison of the primary structures of the A subunits of VT1, VT2 and the ricin A chain revealed three conserved regions (amino acid residues 51-55, 167-171 and 202-207 from the N-terminus of VT1). All three regions of the ricin A chain corresponded in position to the active site of ricin proposed by X-ray crystal diffraction analysis. To determine the relative importance of the conserved amino acid residues for toxin activity of VT1, we prepared VT1 mutants with single amino-acid substitutions by oligonucleotide-directed site-specific mutagenesis. Twenty-two mutants were prepared to examine 14 conserved residues and their cytotoxicities to Vero cells and inhibitory activities on protein synthesis in a rabbit reticulocyte lysate were compared with those of wild-type VT1. Replacement of glutamic acid at position 167 by glutamine and of arginine at position 170 by leucine reduced both activities drastically. These results suggest that glutamic acid at position 167 and arginine at position 170 play the important role in the toxin activity of VT1. A possibility of these mutant toxins to be protective antigen (s) to protect enterohemorrhagic E. Coli infection wil be further studied.
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Report
(3 results)
Research Products
(20 results)
