Grant-in-Aid for international Scientific Research
|Allocation Type||Single-year Grants|
|Research Institution||Tokyo University of Pharmacy & Life Science|
OSHIMA Tairo Tokyo Univ.Pharm & Life Sci., Dept of Molecular Biol., Professor, 生命科学部, 教授 (60167301)
ZAVODSZKY Peter Hungarian Acad.Sci., Inst.Enzymology, 酵素研究所, 教授
PETSKO Gregory Dept.Biochem., Brandeis University, 生化学科, 教授
YAMAGISHI Akihiko Dept.Mol.Biol., Tokyo Univ.Pharm.& Life Sci., 生命科学部, 助教授 (50158086)
TANAKA Nobuo Dept.Life Sci., Tokyo Institute of Technology, 生命理工学部, 教授 (50032024)
ZAVODSZKY Pe ハンガリー科学アカデミー生化学研究所, 教授
PETSKO Grego Brandeis大学, 生化学科, 教授
|Project Period (FY)
1995 – 1996
Completed(Fiscal Year 1996)
|Budget Amount *help
¥3,800,000 (Direct Cost : ¥3,800,000)
Fiscal Year 1996 : ¥1,900,000 (Direct Cost : ¥1,900,000)
Fiscal Year 1995 : ¥1,900,000 (Direct Cost : ¥1,900,000)
|Keywords||Isopropylmalate dehydrogenase / Extreme thermophile / Stabilization of a protein / Crystallography / Chimerie enzyme / 進化分子工学 / イソプロピルリンゴ酸デヒドロゲナーゼ / タンパク質安定性 / タンパク質結晶構造 / 好熱性細菌 / 部位特異的変異 / タンパク質構造機能相関|
An international collaboration between Japan, the United States, and Hungary, has been carried out to clarify the molecular reason why thermophile enzymes are unusually resistant to heat using isopropylmalate dehydrogenase as a model enzyme. The collaboration has been successfully achieved and parts of the results are summarized in international joint papers which will be appeared in scientific journals soon.
1 Molecular design to stabilize chimeric isopropylmalate dehydrogenase between an extreme thermophile (Thermus thermophilus) and a mesophile (Bacillus subtilis) has been done and many stabilized mutants were prepared in the Japanese laboratories. Some of them was distributed among the members.
2 X-ray crystallographic analyzes at low temperature were done in both the US and a Japanese laboratories under close contacts.
3 Molecular cloning of E.coli leuB gene has been done in Japan and distributed to other members. Purification of the enzyme was carried out in Japan by the co-operation of a US and a Japanese researchers. X-ray crystallographic analysis was achieved in the US.Using this cloned gene and 3-D structure, molecular design of the stabilization was done in the US and Hungary.
4 In the Japanese laboratories, the stabilization of the E.coli enzyme has been attempted by using evolutionary molecular engineering techniques.
5 Adaptation of the thermophile enzyme to the mesophilic temperature has been tried in both the US and a Japanese laboratories under close communications. In the Japanese laboratory, an integration-expression system has been constructed and mesophilic temperature-adapted mutants were obtained. Structural analyzes of the mutants are now under way.
6 A researcher from the US visited the Japanese laboratories and joint experiments were carried out. Two Japanese scientists visited the US to discuss the experimental results.