2005 Fiscal Year Final Research Report Summary
Molecular biological analysis of the enzyme networks using the legume isoflavonoid pathway as a model system
| Project/Area Number |
15510183
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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 |
Living organism molecular science
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| Research Institution | Nihon University |
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Principal Investigator |
AKASHI Tomoyoshi Nihon University, Department of Applied Biological Sciences, Assistant, 生物資源科学部, 助手 (80328707)
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| Co-Investigator(Kenkyū-buntansha) |
AYABE Shin-ichi Nihon University, Department of Applied Biological Sciences, Professor, 生物資源科学部, 教授 (40050679)
AOKI Toshio Nihon University, Department of Applied Biological Sciences, Associate Professor, 生物資源科学部, 助教授 (80287606)
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| Project Period (FY) |
2003 – 2005
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| Keywords | Leguminosae / Isoflavonoid / Biosynthesis / Phytoalexin / Enzyme interaction / Glycyrrhiza echinata / Soybean / Lotus japonicus |
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| Research Abstract |
Plants accumulate a variety of natural products. They are biosynthesized by multi-step enzyme reaction processes. Some mechanisms, e.g. enzyme complex, for effective synthesis of specific products in plant cells have been envisaged. Isoflavonoids are biosynthesized mainly in leguminous plants and play important roles in the producer plants. In this study, we cloned cDNAs encoding enzymes of the pathway and attempted to demonstrate the mechanism for the biosynthesis.
cDNAs of 2-hydroxyisoflavanone 4'-O-methyltransferase (HI4'OMT) and 2-hydroxyisoflavanone dehydratase (HID) involved in isoflavone biosynthesis and cDNAs of pterocarpan reductase (PTR) in phytoalexin pathway were cloned from Glycyrrhiza echinata, Lotus japonicus and soybean using functional expression fractionation screenings, RT-PCR and EST information.
Polyketide reductase (PKR) is involved in 6'-deoxychalcone formation. We demonstrated that the recombinant G.echinata PKR protein also mediates the late step of echinatin biosynthesis, i.e.2'-O-methyllicodione reduction. Enzyme recycling of PKR in echinatin biosynthesis is envisaged. Partial purification of G.echinata HI4'OMT was performed using an OMT specific affinity column.HID protein was co-eluted with HI4'OMT, suggesting the protein-protein interaction between HI4'OMT and HID. Finally, to clarify the enzyme function in vivo, experiments incorporating IFS and HID cDNAs in recombinant yeast systems were carried out. The yeast cells expressing both IFS and HID accumulated higher levels of isoflavonoids from exogenously supplied flavanones than the single IFS transformant. It is assumed that cooperation between IFS and HID is required for effective synthesis of isoflavonoids.
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Research Products
(8 results)
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[Journal Article] cDNA cloning and biochemical characterization of S-adenosyl-L-methionine : 2,7,4'-trihydroxyisoflavanone 4'-O-methyltransferase, a critical enzyme of the legume isoflavonoid phytoalexin pathway.2003
Author(s)
Akashi, T., Sawada, Y., Shimada, N., Sakurai, N., Aoki, T., Ayabe, S.
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Journal Title
Plant and Cell Physiology 44(2)
Pages: 103-112
Description
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