Grant-in-Aid for International Scientific Research.
|Section||Joint Research .|
|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)
山岸 明彦 東京薬科大学, 生命科学部, 助教授 (50158086)
田中 信夫 東京工業大学, 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
ZAVODSZKY P. ハンガリー科学アカデミー, 酵素研究所, 教授
PETSKO G. Brandeis大学, 生化学科, 教授
PETSKO Gregory Dept.Biochem., Brandeis University
ZAVODSZKY Peter Hungarian Acad.Sci., Inst.Enzymology
TANAKA Nobuo Dept.Life Sci., Tokyo Institute of Technology
YAMAGISHI Akihiko Dept.Mol.Biol., Tokyo Univ.Pharm.& Life Sci.
|Project Fiscal Year
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 / イソプロプルリンゴ酸デヒドロゲナーゼ / 高度好熱菌 / タンパク質の安定化設計 / 結晶構造解析 / キメラ酵素 / 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.