Multiforn of γ-Glutamyltransfemse in Higher Plants and Cysteine Recycle

高等植物中γ-谷氨酰转移酶的多种形式及半胱氨酸的回收

• 批准号: 14560052
• 负责人: SEKIYA Jiro
• 金额: $ 2.3万
• 依托单位: Kyoto University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2002
• 资助国家: 日本
• 起止时间: 2002 至 2003
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-14560052/
• 关键词: γ-glutamyltransferase; mdish; bound γ-glutamyltransferase; soluble γ-glutamyltransferase; transgenic tobacco; multiform; glutathione catabolism; グルタミル基転移酵素; 可溶性γ-GT; 細胞壁結合性γ-GT

1 Occurrence of two forms of radish γ-glutamyltransferases (GGT; EC.2.3.2.2), soluble and cell wall bound ones, were demonstrated in many plants including radish (Raphanus sativus).2 We cloned three cDNAs (rsggt1, AB098475; rsggt2, AB102676;rsggt3, unregistered) predicted to encode radish soluble GGTs, sequenced them and analyzed their properties. These cDNA seems to encode a heterodimeric type of GGTs homologous to mammalians GGTs.3 These three genes were transcribed by organ and tissue specific manner: rsggt1 was expressed characteristically in immature seeds and cotyledons; rsggt2 in cotyledons, leaves and roots; and rsggt 3 in leaves, roots and immature seeds.4 We transformed tobacco plants with rsggt1 and rsggt2. Transformed tobacco plants grew normally. They developed higher level of GGT activity, although wild type tobacco plants exhibited a very low GGT activity. Expressed GGT activity was found in the bound fraction, although cDNA used encoded soluble radish GGT. To understand this discrepancy remains to be solved in future.5 We purified bound GGTs from radish cotyledons. The bound GGT was a monomeric enzyme unlike soluble GGTs. cDNA was presumed to be highly homologous to Arabidopsis cDNA (AK118068) encoding putative β-1,3-glucanase. Tobacco plants transformed with putative β-1,3-glucanase cDNA exhibited elevated level of GGT activity, but not β-1,3-glucanase.6 We measured thiol compounds in apoplastic space of leaves of transgenic and wild type tobacco plants. Decreased level of glutathione (GSH) and elevated level of cysteinylglycine, a product of GGT reaction, were found in transgenic plants. These findings suggest that cell wall bound GGTs participate in GSH catabolism in apoplastic space.

1 Occurrence of two forms of radish γ-glutamyltransferases (GGT; EC.2.3.2.2), solid and cell wall bound ones, were demonstrated in many plants including radish (Raphanus sativus).2 We cloned three cDNAs (rsggt1, AB098475; rsggt2, AB102676;rsggt3, unregistered) predicted to encode radish soluble GGT，对它们进行了测序并分析其特性。这些cDNA似乎编码了与哺乳动物ggt同源的异二聚体类型的GGTS。3这三个基因以器官和组织的特定方式转录：RSGGT1在未成熟的种子和子叶叶子中表达了特征。 rsggt2在子叶，叶子和根中；和RSGGT 3在叶子，根和未成熟种子中。4我们用RSGGT1和RSGGT2转化了烟草植物。转化的烟草植物正常生长。尽管野生型烟草植物表现出非常低的GGT活性，但他们发展了更高的GGT活性。尽管cDNA使用编码的固体萝卜GGT，但在结合部分中发现了表达的GGT活性。要理解这种差异，将来仍有待解决。5我们从萝卜子叶中纯化了绑定的GGT。与固体GGT不同，结合的GGT是一种单体酶。 cDNA被提出与编码假定的β-1,3-葡聚糖酶的拟南芥cDNA高度同源（AK118068）。烟草植物用推定的β-1,3-葡萄糖酶cDNA转化为GGT活性的升高水平，但β-1,3-葡萄糖酶却没有变化。6我们测量了转基因和野生型烟草植物的叶片中硫醇化合物。在转基因植物中发现了谷胱甘肽（GSH）（GSH）水平（GSH）和胱天但甘氨酸甘氨酸水平的升高。这些发现表明，细胞壁结合GGT参与了质塑料空间中的GSH分解代谢。