Isolation of cDNA clone of key enzymes realting sorbitol synthesis and the analysis of sugar accumulation

山梨醇合成关键酶cDNA克隆的分离及糖积累分析

• 批准号: 04454052
• 负责人: YAMAKI Shohei
• 金额: $ 4.03万
• 依托单位: Nagoya University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (B)
• 财政年份: 1992
• 资助国家: 日本
• 起止时间: 1992 至 1993
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-04454052/
• 关键词: S6PDH; SDH; S6PDH cDNA clone; Sorbitol-6-P dehydrogenase; Sorbitol dehydrogenase; Sorbitol; Sugar accumulation

Both sorbito-6-P dehydrogenase (S6PDH) and sotbitol dehydrogenase (SDH) are important as key enzymes to accumulate sugar into fruit tissue of apple. The purpose of this research is to make clear the role of lthese enzymes by purifying and characterizing them, and to clarify the regulation on gene expression of S6PDH by cloning the cDNA.We cucceeded to isolate both enzymes through some purification steps. Especially, the isolation of S6PDH is very worth because this was firstly isolated from plant tissues. From the properties of purified enzymes, S6PDH was certified to react toward S6P synthesis and SDH toward the convertion of sorbitol into fructose in vivo. Further, we succeeded to isolate the cDNA clone of S6PDH using apple seedling. The cDNA is composed of a open reading fram of 930 bases coding 310 mino acid. The molecular weight of S6PDH was 35,000 and corresponded to the molecular weight estimated by SDS-PAGE.Next, the gene expression of S6PDH with germination of apple seeds was studied using this cDNA probe. It was suggested that the gene expression of S6PDH activity acts under some post-transcriptional or translational regulations since m-RNA of S6PDH appeared 2 days ealier than the protein synthesis of S6PDH.Infuture, it will be expected to make clear the detail mechanism of sugar accumulation into fruit throught the study of gene expression of SDH by cloning the cDNA.

山梨醇脱氢酶（SDH）和山梨醇-6-磷酸脱氢酶（S6 PDH）是苹果果实组织糖积累的关键酶。本研究的目的是通过纯化和鉴定这两种酶来阐明它们的作用，并通过克隆S6 PDH的cDNA来阐明它们对S6 PDH基因表达的调控。特别是S6 PDH的分离是非常有价值的，因为这是首次从植物组织中分离到的。从纯化的酶的性质，S6 PDH被证明是对S6 P的合成和SDH对山梨醇转化为果糖在体内的反应。利用苹果幼苗成功地克隆了S6 PDH的cDNA克隆。该cDNA由930个碱基组成的开放阅读弗拉姆，编码310个氨基酸。S6 PDH的分子量为35，000，与SDS-PAGE测定的分子量一致。S6 PDH mRNA的表达比S6 PDH蛋白质合成早2 d，说明S6 PDH活性基因的表达受转录后或翻译调控。

项目成果

金山喜則: "果実が甘くなるしくみ" 化学と生物. 31. 578-587 (1993)

Yoshinori Kanayama：“水果如何变甜”化学与生物学 31. 578-587 (1993)。

Y.Kanayama: "Purification and properties of NADP-dependent sorbitol-6-phosphate dehydrogenase from apple seedlings" Plant Cell Physiol.34. 819-823 (1993)

Y.Kanayama：“苹果幼苗中 NADP 依赖性山梨醇 6-磷酸脱氢酶的纯化和特性”Plant Cell Physiol.34。

T.Moriguchi: "Levels and role of sucrose synthase,sucrose-phosphate synthase,and acid invertase in sucrose accumulation in fruit of Asian pear" J.Amer.Soc.Hort.Sci.117. 274-278 (1992)

T.Moriguchi：“蔗糖合酶、蔗糖磷酸合酶和酸性转化酶在亚洲梨果实蔗糖积累中的水平和作用”J.Amer.Soc.Hort.Sci.117。

Y.Kanayama and S.Yamaki: "Mechanism o sugar accumulation in fruit" KASEAA. 31. 578-586 (1993)

Y.Kanayama 和 S.Yamaki：“水果中糖积累的机制”KASEAA。

Y.Kanayama: "Nucleotide sequence of a cDNA encoding NADP-sorbitol-6-phosphate dehydrogenase from apple" Plant Physiol.100. 1607-1608 (1992)

Y.Kanayama：“编码来自苹果的 NADP-山梨糖醇-6-磷酸脱氢酶的 cDNA 的核苷酸序列”Plant Physiol.100。