| Project/Area Number |
13480206
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Functional biochemistry
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| Research Institution | The University of Tokyo |
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Principal Investigator |
ITO Koichi The University of Tokyo, Institute of medical science, Assistant professor, 医科学研究所, 助手 (10262073)
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| Co-Investigator(Kenkyū-buntansha) |
NAKAMURA Yoshikazu The University of Tokyo, Institute of medical science, Professor, 医科学研究所, 教授 (40114590)
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| Project Period (FY) |
2001 – 2002
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| Project Status |
Completed (Fiscal Year 2002)
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| Budget Amount *help |
¥15,200,000 (Direct Cost: ¥15,200,000)
Fiscal Year 2002: ¥5,100,000 (Direct Cost: ¥5,100,000)
Fiscal Year 2001: ¥10,100,000 (Direct Cost: ¥10,100,000)
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| Keywords | TRANSLATION TERMINATION / TRANSLATIONAL REGULATION / RIBOSOME / RECODING / RELEASE FACTOR / PROTEIN SYNTHESIS / TRNA-PROTEIN MIMICRY / REGULATION OF GEHE EXPRESSION / 翻訳抑制 / ペプチド鎖解離因子 |
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| Research Abstract |
A yeast eRF1-domain swap containing Tetrahymena domain 1 responded only to UGA in vitro and failed to complement a defect in yeast eRF1 in vivo at 37 degrees C. This finding demonstrates that decoding specificity of eRF1 from variant code organisms resides at domain 1. However, the wild-type eRF1 hybrid fully restored the growth of eRF1-deficient yeast at 30 degrees C. Tetrahymena eRF1 contains a variant sequence, KATNIKD, at the tip of domain 1. The TASNIKD variant of hybrid eRF1 rendered the eRF1-nullified yeast viable, although in an in vitro assay, the same hybrid eRF1 responded only to UGA. Nevertheless, the yeast eRF1 bearing the KATNIKD motif instead of the TASNIKS heptapeptide present in higher eukaryotes remains omnipotent in vivo. Collectively, these data suggest that variant genetic code organisms like Tetrahymena have an intrinsic potential to decode three stop codons in vivo, and that interaction within domain 1 between the KAT tripeptide and other sequences modulates the decoding specificity of Tetrahymena eRF1.
Thermus thermophilus RRF is nonfunctional in Escherichia coli. It became functional upon introducing a plasmid expressing E. coli EF-G with surface changes in its tRNA-mimic domain or by replacing the E. coli EF-G tRNA-mimic domain by the Thermus domain. Thermus RRF could also be activated by introducing surface substitutions in its anticodon arm-mimic region. These gain-of-function phenotypes depend on the combination of heterologous EF-G and RRF alleles. These mutational studies suggest that EF-G motor action is transmitted to RBF by specific surface contacts between the domains that mimic the anticodon arm.
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