Design and synthesis of hybrid-type molecular probes useful for elucidation of molecular mechanisms of physiological events caused by brassinosteroids

杂合型分子探针的设计和合成有助于阐明油菜素类固醇引起的生理事件的分子机制

• 批准号: 15590034
• 负责人: SETO Hideharu
• 金额: $ 2.05万
• 依托单位: RIKEN
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2003
• 资助国家: 日本
• 起止时间: 2003 至 2004
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-15590034/
• 关键词: plant hormone; brassinosteroid; synthesis; hybrid-type probe; photoaffinity probe; receptor; biotin; phenyldiazirine; 光アフィニティー標識; ビオチン標識; 水酸化反応; methyl(trifluoromethyl)dioxirane

Brassinosteroids(BRs) constitute a class of steroidal plant hormones, which are essential for plant growth and development. Many efforts have been made in a wide range of research fields in order to elucidate the real features of BR actions at molecular level in plants, using technologies based on molecular biology. In this project, I aimed the design and synthesis of BR-based hybrid-type molecular probes useful for elucidation of molecular mechanisms of physiological events caused by BRs. and made considerable achievements as follows.1.Starting with natural BRs, brassinolide(BL) and castasterone(CS), the synthesis of several BR-based hybrid-type molecular probes : i.e., phenyldiazirine/biotin double-labelled BR, biotin labeled BR and dexamethazone labeled BR, were completed.2.In early works on BR perception mechanism in plant, we elucidated that a transmembrane receptor kinase identified from Arabidopsis, BRI1,is a receptor of BRs. The photoaffinity labeling experiments using aryldiazirine/biotin double-labelled CS synthesized as a photoaffinity probe clarified that BR directly binds to BRI1 without other proteins or peptides. Furthermore, the minimal binding domain for BR consisting of 94 amino acids was identified, which was a new binding domain for steroid hormones.3.26-hydroxy-CS and -BL synthesized in this project as synthetic intermediates to BR-based hybrid-type molecular probes played a crucial role in the functional analysis of the Arabidopsis CYP72B1,BAS1 and SOB7 genes. It was elucidated that these genes inactivated BRs through 26-hydroxylation of active BRs, CS and BL.4.In connection with this project, four positional isomers of mono-O-β-D-glucopyranosides of BL and CS with tetrahydroxy groups were prepared. These glycosides played a crucial role in the functional analysis of the Arabidopsis UGT73C5 gene expressing a UPD-glycosyltransferase, UGT73C5. It was elucidated that UGT73C5 specifically glycosylated 23-hydroxyl function of CS and BL.

类甾醇（BRs）是一类甾体植物激素，对植物的生长发育具有重要作用.为了在分子水平上阐明BR在植物中作用的真实的特征，人们利用基于分子生物学的技术在广泛的研究领域中进行了许多努力。本研究的目的是设计和合成基于BR的杂交型分子探针，用于阐明BR引起的生理事件的分子机制。1.从天然BR、油菜素内酯（BL）和Castasterone（CS）出发，合成了几种BR杂交型分子探针，2.在植物BR感受机制的早期研究中，我们发现拟南芥中存在一种跨膜受体激酶BRI 1，它是BR的受体。利用合成的芳基二氮杂环丙烯/生物素双标记CS作为光亲和探针进行光亲和标记实验，澄清了BR直接结合BRI 1而不需要其他蛋白或肽。此外，还鉴定了BR的最小结合域，该结合域由94个氨基酸组成，是一个新的类固醇激素结合域。本项目合成的3，26-hydroxy-CS和-BL作为BR杂交型分子探针的合成中间体，在拟南芥CYP 72 B1、BAS 1和SOB 7基因的功能分析中发挥了重要作用。4.结合本课题，合成了4个含四羟基的BL和CS单-O-β-D-吡喃葡萄糖苷的位置异构体。这些糖苷在表达β-糖基转移酶UGT 73 C5的拟南芥UGT 73 C5基因的功能分析中发挥了至关重要的作用。已阐明UGT 73 C5特异性糖基化CS和BL的23-羟基功能。

项目成果

植物の生長ホルモン・ブラシノステロイドと受容体BRI1の結合メカニズム

植物生长激素油菜素内酯与受体BRI1的结合机制

• 发表时间: 2005
• 期刊: BRAIN Techno News 109(印刷中)
• 作者: 瀬戸秀春;木下俊則
• 通讯作者: 木下俊則

Brassinosteroids bind to the extracellular domainof plant receptor kinase BRI1.

油菜素类固醇与植物受体激酶 BRI1 的胞外域结合。

• 发表时间: 2005
• 期刊: Cell Technology (in Japanese) 433
• 作者: T.Kinoshita;H.Seto
• 通讯作者: H.Seto

CYP72B1 inactivates brassinosteroid hormones : intersection between photomorphogenesis and plant steroid signal transduction

CYP72B1 灭活油菜素类固醇激素：光形态发生与植物类固醇信号转导之间的交叉

• 发表时间: 2003
• 期刊: Plant Physiol. 133
• 作者: T.Kinoshita;A.Cano-Delgado;H.Seto;S.Hiranuma;S.Fujioka;S.Yoshida;J.Chory;E.M.Turk et al.
• 通讯作者: E.M.Turk et al.

CYP72B1 inactivates brassinosteroid hormones : intersection between photomorphogenesis and plant steroid signal transduction.

CYP72B1 使油菜素类固醇激素失活：光形态发生和植物类固醇信号转导之间的交叉。

• 发表时间: 2003
• 期刊: Plant Physiol. 133
• 作者: T.Kinoshita;A.Cano-Delgado;H.Seto;S.Hiranuma;S.Fujioka;S.Yoshida;J.Chory;E.M.Turk et al.;E.M.Turk et al.
• 通讯作者: E.M.Turk et al.

ブラシノステロイドは植物受容体キナーゼBRI1の細胞外ドメインに結合する

油菜素类固醇与植物受体激酶 BRI1 的胞外域结合

• 发表时间: 2005
• 期刊: 植物細胞工学 24
• 作者: 木下俊則;瀬戸秀春
• 通讯作者: 瀬戸秀春