Elucidation of the substrate supplying system in Toxoplasma gondii specific GTP-PK
| Project/Area Number |
23590494
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Multi-year Fund |
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| Section | 一般 |
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| Research Field |
Parasitology (including Sanitary zoology)
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| Research Institution | Keio University |
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Principal Investigator |
ASAI TAKASHI 慶應義塾大学, 医学部, 講師 (30159355)
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| Co-Investigator(Kenkyū-buntansha) |
IMADA Mihoko 慶應義塾大学, 医学部, 助教 (50445201)
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| Project Period (FY) |
2011 – 2013
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| Project Status |
Completed (Fiscal Year 2013)
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| Budget Amount *help |
¥5,200,000 (Direct Cost: ¥4,000,000、Indirect Cost: ¥1,200,000)
Fiscal Year 2013: ¥1,040,000 (Direct Cost: ¥800,000、Indirect Cost: ¥240,000)
Fiscal Year 2012: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
Fiscal Year 2011: ¥2,210,000 (Direct Cost: ¥1,700,000、Indirect Cost: ¥510,000)
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| Keywords | トキソプラズマ / ピルビン酸キナーゼII / ホスホエノールピルビン酸カルボキシキナーゼ / GDP-PK / PEPCK / 寄生原虫 / ミトコンドリア / ピルビン酸キナーゼ / 大腸菌での発現 / アピコプラスト / 細胞内小器官での局在 |
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| Research Abstract |
A pyruvate kinase II in mitochondria and apicoplast of Toxoplasma gondii has not been known its physiological function. To understand the physiological function of this enzyme, the substrate supplying system of this enzyme was investigated. For the first time, a production of two recombinant PEPCKs was studied. The genes of enzymes were produced by PCR using a cDNA of RH strain of T. gondii and reconstituted into a protein producing plasmid pGEX. Type I gene of PEPCK was recognized, however, type II gene was completely recognized. After several trials to other protein producing plasmid (pET etc.), we found that pCold TF plasmid producing a protein at low temperature could produce the type I recombinant PEPCK as a GST-fusion protein. Since type II PEPCK gene could not been obtained by PCR, we tried several PCR techniques (Taq polymerase, annealing Tm, thermal cycler, etc.). Finally, type II gene was obtained.
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Report
(4 results)
Research Products
(23 results)
