| Project/Area Number |
11470492
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Biological pharmacy
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| Research Institution | Riken |
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Principal Investigator |
TSUJIMOTO Masafumi RIKEN,DEPT.OF CELL BIOCHEM, CHIEF SCIENTIST, 細胞生化学研究室, 主任研究員 (00281668)
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| Co-Investigator(Kenkyū-buntansha) |
NOMURA Yosihiro TOKYO UNIV. AGRI. TECH, AGRL, RESERCH ASSCIATE, 農学部, 助手 (10228372)
MATSUMOTO Hideo RIKEN, DEPT.OF CELL BIOCHEM, POSTDOCTRAL FELLOW, 細胞生化学研究室, 基礎科学特別研究員 (00312257)
HATTORI Akira RIKEN, DEPT.OF CELL BIOCHEM, STAFF SCIENTIST, 細胞生化学研究室, 研究員 (50300893)
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| Project Period (FY) |
1999 – 2001
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| Project Status |
Completed (Fiscal Year 2001)
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| Budget Amount *help |
¥6,500,000 (Direct Cost: ¥6,500,000)
Fiscal Year 2001: ¥2,600,000 (Direct Cost: ¥2,600,000)
Fiscal Year 2000: ¥3,900,000 (Direct Cost: ¥3,900,000)
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| Keywords | AMINOPEPTIDASE / OXYTOCINASE / PLACENTAL LEUCINE AMINOPEPTIDASE / ADIPOCYTE-DERIVED LEUCINE AMINBOPEPTIDASE / BIOACTIVE PEPTIDE / VASOPRESSIN / BLOOD PRESSURE / cDNA CLONING / 尿細管 / プロモーター領域 / 神経細胞 / ペプチドホルモン / CHO細胞 / タンパク質結晶化 / 金属酵素 / 胎盤性ロイシンアミノペプチターゼ |
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| Research Abstract |
It is well known that placental leucine aminopeptidase (P-LAP)/oxytocinase increases in matemal serum during pregnancy and maintaihs normal pregnancy by preventing the action of oxytocin. We have cloned the cDNA encoding P-LAP for the first time and shown that the enzyme is a type II membrane- spanning protein. In this study we have further analysed pathological/physiological properties of the eazyme in detail.
We have constructed a large-scale production system of recombinant P-LAP employing CHO cells and analysed the enzymatic properties of the enzyme. It was found that in addition to oxytocin and vasopressin, the enayme degraded some neuronal peptides such as Met-enkephalin and dynorphin A. Since the eazyme was expressed in neuronal cells and increased in its expression during neuronal differentiation, it is considered that P-LAP plays an important role in the regulation of neuronal cells in the brain. Since antidiuretic hormone vasopressin, a good substrate for P-LAP, regulates wate
… More
r reabsorption in renal collecting duct principal cells and expression of P-LAP in the kidne was detected, we have analysed the effect of the hormone on the translocation of the eazyme rom intracellular vesicle to plasma membrane. It was found that after treatment of NRK52E cells with vasopressin, rapid translocation was clearly detected. These results suggested the existence of the P-LAP mediated negative feedback mechabism of the vasopressin action in the kidney. Moreover we have analysed the promoter function of P-LAP gene and found that two
transcription factors, AP-2 and ikaros, were important for the regulation of the gene.
By searching a database, we have cloned a novel enzyme tenned adipocyte-derived leucine aminopeptidase (A-LAP). Detailed structural analysis indicated that P-LAP and A-LAP should be classified into oxyiocinase sub-fatnily. Enzymatic analysis after construcdon of a large-scale production system of recombinant A-LAP suggested that the enzyme played a role in the regulation of blood pressure through the inactivation of angiotensin 11 and/or the generation of bradyki' nin in the kidney., Less
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