リグニン オヨビ リグナン セイゴウセイ ニ カンヨスル O - メチルトランスフェラーゼ オヨビ ピノレジノール レダクターゼ ノ トクセイ カイメイ

京都大学0048新制・課程博士博士(農学)甲第14173号農博第1734号新制||農||964(附属図書館)学位論文||H20||N4412(農学部図書室)UT51-2008-N490京都大学大学院農学研究科応用生命科学専攻(主査)教授 梅澤 俊明, 教授 宮川 恒, 教授 矢﨑 一史学位規則第4条第1項該当Doctor of Agricultural ScienceKyoto UniversityDA

Basic Studies towards Elucidation of Heartwood Formation Mechanisms

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Characterization of Arabidopsis thaliana Pinoresinol Reductase, a New Type of Enzyme Involved in Lignan Biosynthesis*

A lignan, lariciresinol, was isolated from Arabidopsis thaliana, the most widely used model plant in plant bioscience sectors, for the first time. In the A. thaliana genome database, there are two genes (At1g32100 and At4g13660) that are annotated as pinoresinol/lariciresinol reductase (PLR). The recombinant AtPLRs showed strict substrate preference toward pinoresinol but only weak or no activity toward lariciresinol, which is in sharp contrast to conventional PLRs of other plants that can reduce both pinoresinol and lariciresinol efficiently to lariciresinol and secoisolariciresinol, respectively. Therefore, we renamed AtPLRs as A. thaliana pinoresinol reductases (AtPrRs). The recombinant AtPrR2 encoded by At4g13660 reduced only (–)-pinoresinol to (–)-lariciresinol and not (+)-pinoresinol in the presence of NADPH. This enantiomeric selectivity accords with that of other PLRs of other plants so far reported, which can reduce one of the enantiomers selectively, whatever the preferential enantiomer. In sharp contrast, AtPrR1 encoded by At1g32100 reduced both (+)- and (–)-pinoresinols to (+)- and (–)-lariciresinols efficiently with comparative kcat/Km values. Analysis of lignans and spatiotemporal expression of AtPrR1 and AtPrR2 in their functionally deficient A. thaliana mutants and wild type indicated that both genes are involved in lariciresinol biosynthesis. In addition, the analysis of the enantiomeric compositions of lariciresinol isolated from the mutants and wild type showed that PrRs together with a dirigent protein(s) are involved in the enantiomeric control in lignan biosynthesis. Furthermore, it was demonstrated conclusively for the first time that differential expression of PrR isoforms that have distinct selectivities of substrate enantiomers can determine enantiomeric compositions of the product, lariciresinol