| Project/Area Number |
02404015
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| Research Category |
Grant-in-Aid for General Scientific Research (A)
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| Allocation Type | Single-year Grants |
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| Research Field |
林産学
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| Research Institution | KYOTO UNIVERSITY |
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Principal Investigator |
SHIMADA Mikio (1991-1992) Wood Res. Inst., Kyoto Univ Professor, 木質科学研究所, 教授 (50027166)
樋口 隆昌 (1990) 京都大学, 木材研究所, 教授 (00027153)
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| Co-Investigator(Kenkyū-buntansha) |
UMEZAWA Toshiaki Wood Res. Inst., Kyoto Univ.,Instructor, 木質科学研究所, 助手 (80151926)
KURODA Hiroyuki Wood Res. Inst., Kyoto Univ.,Instructor, 木質科学研究所, 助手 (00115841)
島田 幹夫 京都大学木材研究所, 講師 (50027166)
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| Project Period (FY) |
1990 – 1992
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| Project Status |
Completed (Fiscal Year 1992)
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| Budget Amount *help |
¥25,900,000 (Direct Cost: ¥25,900,000)
Fiscal Year 1992: ¥1,200,000 (Direct Cost: ¥1,200,000)
Fiscal Year 1991: ¥3,200,000 (Direct Cost: ¥3,200,000)
Fiscal Year 1990: ¥21,500,000 (Direct Cost: ¥21,500,000)
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| Keywords | Enantioselectivity / (-)-Secoisolariciresinol / Lariciresinol / Callus / Forsythia sp / Lignan synthase gene / PAL遺伝子 / リグニン / リグナン / 生合成 / エナンチオ選択的カップリング / キラルHPLC / リグナン生合成酵素遺伝子 / 立体選択的カップリング / PCR |
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| Research Abstract |
First, systems for chiral HPLC separation and chiral LC-MS analysis of several lignans were established, which provide important basis for tracer experiments with stable isotopes in studies of lignan biosynthesis. The chiral LC-MS system was successfully applied to the assay of enzyme activity for enantioselective formation of (-)-secoisolariciresinol.
Second, callus was successfully induced from explants of Forsythia koreana and the production systems for the cultured cells producing both lignans and lignan-synthesizing enzymes have been established. The calli have shown to produce several lignan glycosides. Many of the calli produced significant amounts of matairesinol (aglycones), exhibiting enzymatic activity forming secoisolariciresinol, a precursor of matairesinol biosynthesis, from coniferyl alcohol; comparable to those of field growing F.koreana plants. The results indicate potential use of F.koreana tissue cultures as a source of lignan synthesizing enzyme(s).
Cell-free extracts from F.koreana catalyzed enantioselective formation of (-)-, but not (+)-,[^2H_<10>]secoisolariciresinol from [^2H_<10>]- coniferyl alcohol in the presence of NADPH and H_2O_2. Surprisingly, (-)- pinoresinol and (-)-lariciresinol were also formed in the incubation although they are not natural enantiomers isolated from F.koreana. In addition, the enzyme system catalyzed enantioselective reduction of (+)- lariciresinol to afford (-)-secoisolariciresinol in the presence of NADPH. Mechanisms for the enantioselective formation of (-)-secoisolariciresinol from coniferyl alcohol were proposed. The optical activity of lignans is formed probably by some of peroxidases and NADPH:quinone oxidoreductases. There are multiple forms of those enzymes in the young shoots from F. koreana. We also discussed strategies on the molecular cloning for the enzymes involved in lignan biosynthesis.
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