| Project/Area Number |
11660068
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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 |
応用微生物学・応用生物化学
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| Research Institution | HOKKAIDO UNIVERSITY |
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Principal Investigator |
ITO Hiroyuki Hokkaido Univ., Grad.School of Agr., Associate Prof., 大学院・農学研究科, 助教授 (10241366)
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| Project Period (FY) |
1999 – 2000
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| Project Status |
Completed (Fiscal Year 2000)
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| Budget Amount *help |
¥3,100,000 (Direct Cost: ¥3,100,000)
Fiscal Year 2000: ¥1,700,000 (Direct Cost: ¥1,700,000)
Fiscal Year 1999: ¥1,400,000 (Direct Cost: ¥1,400,000)
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| Keywords | Starch biosynthesis / Allosteric regulation / ADPglucose pyrophosphorylase / Transgenic plants / Mutated enzymes |
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| Research Abstract |
Plant ADPglucose pyrophosphorylase (AGPase) catalyzes the supply of ADPglucose that is a sole substrate of starch biosynthesis. AGPase is a heterotetrameric enzyme composed of two large and two small subunits, and allosterically regulated by 3-phosphoglycerate and Pi. The aim in this study was a quantitative alteration of starch contents by improvements of allosterical properties of AGPase.
1) Mutant AGPases with low sensitivities to allosteric effectors were prepared from E.coli expressing the native samll subunit cDNA and the mutated large subunit cDNA of Arabidopsis AGPase.
2) The ApL1 gene conaining the promoter region was isolated from an Arabidopsis genomic library. The sequence revealed that the gene consists of 14 introns and 15 exons.
3) Based on the results of mutant AGPases prepared from E.coli cells, the same mutations were introduced in the ApL1 gene by a site-directed mutagenesis. The mutant genes were ligated under the self promoter. Arabidopsis TL46 plants, that are deficient in ApL 1 protein in leaves, were transformed with the constructed mutant genes.
4) In leaves of transgenic plants expressing allosteric mutant ApL1, the maximum starch accumulation was more than 4-fold of TL46 plants and about 1.2-fold of wild type plants.
These results suggest that the allosteric properties of AGPase can alter the rate of starch biosynthesis.
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