Molecular pathological analysis of inborn error of metabolism with mineralization defects
| Project/Area Number |
16591854
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Functional basic dentistry
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| Research Institution | Niigata University |
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Principal Investigator |
ODA Kimimitsu Niigata University, Institute of Medicine and Dentistry, professor, 医歯学系, 教授 (10122681)
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| Co-Investigator(Kenkyū-buntansha) |
AMAYA Yoshihiro Niigata University, Institute of Medicine and Dentistry, associate professor, 医歯学系, 助教授 (50193032)
AMIZUKA Norio Niigata University, Center for Transdiciplinary Research, Professor, 超域研究機構, 教授 (30242431)
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| Project Period (FY) |
2004 – 2005
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| Project Status |
Completed (Fiscal Year 2005)
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| Budget Amount *help |
¥3,300,000 (Direct Cost: ¥3,300,000)
Fiscal Year 2005: ¥1,200,000 (Direct Cost: ¥1,200,000)
Fiscal Year 2004: ¥2,100,000 (Direct Cost: ¥2,100,000)
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| Keywords | hypophsphatasia / alkaline phosphatase / frameshift-mutation / inborn error of metabolism / ubiquitination / proteasome / dominant mode of transmission / mineralization / 先天性骨代謝異常 / 優性遺伝 / Tet-On細胞 / 分解 |
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| Research Abstract |
Hypophosphatase is a genetic disease characterized by defective mineralization of bone and tooth and reduction in serum alkaline phosphatase activity. Various mutations in the tissue-nonspecific alkaline phosphatase gene (TNSALP) are responsible for a wide variety of clinical symptoms of the disease. So far 178 mutations have been reported, though little is known about the molecular basis of this disease. We have been studying effects of several missense mutations on the biosynthesis and catalytic function of TNSALP molecule in mammalian cells expressing each TNSALP mutant. The aim of this research was conducted to elucidate the effect of two mutations : a frame shift caused by a thymidine deletion at 1559 of cDNA (1559delT) and replacement of alanine with threonine at position 99 of TNSALP polypeptide (A99T).
1. 1559delT : In combination with an in vitro translational system, we confirmed that 1559delT is synthesized as a larger polypeptide with 80 amino acid residues at C-terminus. Due to this extra amino acids, 1559delT is no longer a membrane bound enzyme via GPI (glycosylphosphatidyl-inositol), but a soluble enzyme possessing alkaline phosphatase activity. However, since majority of newly synthesized 1559delT forms a disulfide-bonded high-molecular-mass aggregate in the ER, undergo polyubiquitination and is degraded in the proteasome, only a small portion of the 1559delT was secreted into the medium.
2. A99T : Like wild-type TNSALP, A99T was found to expressed as a 80 kDa mature form on the cell surface via GPI, though in contrast to the wild type, though A99T exhibited no enzyme activity. This indicates that alanine at position 99 is involved in catalytic function of TNSALP. Importantly, A99T was found to form a heterodimer with the wild-type polypeptide when coexpressed with the wild type, presumably accounting for its dominant mode of transmission.
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Report
(3 results)
Research Products
(18 results)
