Incorpoation of biocitin into a protein by expanding the genetic code
| Project/Area Number |
24710231
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| Research Category |
Grant-in-Aid for Young Scientists (B)
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| Allocation Type | Multi-year Fund |
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| Research Field |
System genome science
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| Research Institution | Tokyo University of Science |
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Principal Investigator |
UMEHARA Takuya 東京理科大学, 基礎工学部, 助教 (00415548)
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| Research Collaborator |
YANAGISAWA Tatsuo
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| Project Period (FY) |
2012-04-01 – 2015-03-31
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| Project Status |
Completed (Fiscal Year 2014)
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| Budget Amount *help |
¥4,550,000 (Direct Cost: ¥3,500,000、Indirect Cost: ¥1,050,000)
Fiscal Year 2014: ¥1,170,000 (Direct Cost: ¥900,000、Indirect Cost: ¥270,000)
Fiscal Year 2013: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000)
Fiscal Year 2012: ¥1,820,000 (Direct Cost: ¥1,400,000、Indirect Cost: ¥420,000)
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| Keywords | 非天然アミノ酸 / タンパク質翻訳 / tRNA / ピロリジン |
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| Outline of Final Research Achievements |
Since pyrrolysly-tRNA synthetase (PylRS)/pyrrolysine tRNA (tRNA(Pyl)) pair is orthogonal to aminoacyl-tRNA synthetase/tRNA pairs in various organisms, PylRS/tRNA(Pyl) pair has been used as one of the parental molecules to cotranslatinally incorporate an unnatural amino acid into a protein. In this project, I tried to develop the system to produce a biotinylated protein in E. coli by using PylRS/tRNA(Pyl) pair and biotinyl-L-lysine (biocitin). Although several challenges of in vivo selection to PylRS libraries with saturated mutations, many PylRS mutants recognizing natural amino acid were isolated as higher background. In addition to the selection, I constructed the PylRS mutant based on the docking model of PylRS and biocitin, which however showed the activity with very little. These results suggest that large modification of the active site in PylRS is required to obtain PylRS mutant that can charge biocitin onto tRNA(Pyl).
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Report
(4 results)
Research Products
(29 results)
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[Journal Article] Glycols modulate terminator stem stability and ligand-dependency of a glycine riboswitch2013
Author(s)
Hamachi, K., Hayashi, H., Shimamura, M., Yamaji, Y., Kaneko, A., Fujisawa, A., Umehara, T. and Tamura, K
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Journal Title
BioSystems
Volume: 113
Issue: 2
Pages: 59-65
DOI
Related Report
Peer Reviewed
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[Journal Article] RNA tetraplex as a primordial peptide synthesis scaffold2012
Author(s)
Umehara, T., Kitagawa, T., Nakazawa, Y., Yoshino, H., Nemoto, R. and Tamura, K
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Journal Title
BioSystems
Volume: 109
Issue: 2
Pages: 145-150
DOI
Related Report
Peer Reviewed
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