2000 Fiscal Year Final Research Report Summary
Functional alteration of cereal starches using the genes encoding starch-synthesizing enzymes
| Project/Area Number |
10556029
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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 | 展開研究 |
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| Research Field |
食品科学・栄養科学
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| Research Institution | University of Tsukuba |
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Principal Investigator |
BABA Tadashi University of Tsukuba, Institute of Applied Biochemistry, Professor, 応用生物化学系, 教授 (40165056)
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| Co-Investigator(Kenkyū-buntansha) |
MATSUI Hirokazu University of Hokkaido, Graduate School of Agriculture, Professor, 大学院・農学研究科, 教授 (90109504)
FUJIMURA Tatsuhito University of Tsukuba, Institute of Agricultural and Forest Engineering, Professor, 農林工学系, 教授 (70292513)
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| Project Period (FY) |
1998 – 2000
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| Keywords | starch / amylose / amylopectin / rice / branching enzyme / starch synthase / gene / developing seeds |
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| Research Abstract |
To examine whether the genes encoding starch-synthesizing enzymes are able to utilize alteration of the properties of cereal starches, we have carried out biochemical experiments using these genes, and have obtained the following results :
1) Three isoforms of starch branching enzyme, termed RBE1, RBE3, and RBE4, were purified from crude extracts of rice developing seeds. On the basis of the amino-terminal amino acid sequences of the purified proteins, we have identified and characterized cDNA clones encoding each of RBE1, RBE3, and RBE4. An expression system for production of recombinant enzymes in soluble extracts of Escherichia coli cells has been established using the cDNA fragments of RBE1, RBE3, and RBE4.
2) Purified recombinant RBE1 was capable of branching amylose and amylopectin more rapidly than recombinant RBE3 and RBE4. The branched α-glucans produced by the recombinant enzymes from potato amylose revealed the different patterns of oligosaccharide chain transfer ; RBE1 predominantly transferred longer chains of more than 10 glucose units, whereas RBE3 preferred to transfer short chains of 6 and 7 glucose units. RBE4 showed both characteristics of RBE1 and RBE3 in the transferability of oligosaccharide chains.
3) Northern blot analysis demonstrated that the RBE4 gene is expressed in both leaves and developing seeds. The RBE4 gene was distinguished from the RBE1 and RBE3 genes by expression at the earlier stages of seed development. It is thus possible that RBE4 may play an important role in the early stages of amylopectin synthesis.
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