| Project/Area Number |
16K18689
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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 |
Applied biochemistry
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| Research Institution | Tokai University |
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Principal Investigator |
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| Project Period (FY) |
2016-04-01 – 2018-03-31
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| Project Status |
Completed (Fiscal Year 2017)
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| Budget Amount *help |
¥4,160,000 (Direct Cost: ¥3,200,000、Indirect Cost: ¥960,000)
Fiscal Year 2017: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
Fiscal Year 2016: ¥2,210,000 (Direct Cost: ¥1,700,000、Indirect Cost: ¥510,000)
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| Keywords | L-スレオニン酸脱水素酵素 / 酵素化学 / 結晶構造解析 / 植物疫病菌 / L-スレオニン脱水素酵素 / NAD(P) / 選択性農薬 / Ｘ線結晶構造解析 |
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| Outline of Final Research Achievements |
A gene encoding an L-Threonine dehydrogenase (ThrDH; PITG_05140) homologue was identified in the Phytophthora infestans. After transforming E. coli with pET-PITG_05140, an expression vector harboring the gene, the transformant cells exhibited a high level of L-ThrDH activity, and the enzyme was readily purified from the cells’ crude extract in one simple step: Talon metal affinity chromatography. The purified enzyme showed a single protein band on SDS-PAGE, and its N-terminal sequence was determined to be SDKIIHLTDD. SDS-PAGE showed the subunit molecular mass of P. infestans ThrDH to be about 55 kDa, which is consistent with the molecular weight calculated from the amino acid sequence. Non-tagged P. infestans ThrDH acted on L-threonine and DL-3-hydroxynorvaline, and the enzyme was partially inhibited by L-cysteine and N-acetylglycine.
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