Study of tissue-specific dihydrodiol dehydrogenase

组织特异性二氢二醇脱氢酶的研究

• 批准号: 11672175
• 负责人: HARA Akira
• 金额: $ 2.3万
• 依托单位: Gifu Pharmaceutical University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1999
• 资助国家: 日本
• 起止时间: 1999 至 2000
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11672175/
• 关键词: dihydrodiol dehydrogenase; D-xylose dehydrogenase; new protein family; structure-function relationship; site-directed mutagenesis; anti-inflammatory drug; gene structure; 新規蛋白質ファミリー

Dihydrodiol dehydrogenase (DD, EC 1.3.1.20), which catalyzes the NADP-dependent oxidation of trans-dihydrodiols of aromatic hydrocarbons, has been shown to be involved in cytotoxicity of the aromatic hydrocarbons. The enzyme exits in multiple forms in mammalian tissues. Of the multiple forms dimeric DD is tissue-specifically expressed, but its structure and physiological function have not been elucidated. This study was performed to elucidate the structure of dimeric DD, its relationship to the function and possible clinical significance. The results obtained are summarized.1. Dimeric DD in monkey kidney, pig and dog liver was demonstrated to be identical with NADP-dependent D-xylose dehydrogenase (EC 1.1.1.179).2. The protein sequencing and/or cDNA cloning primary structures of dimeric DDs in rabbit lens, human intestine and the above animal tissues revealed that the enzymes constitute a new protein family with prokaryotic proteins including glucose-fructose oxidoreductase.3. The site … More -directed mutagenesis on the monkey dimeric DD suggested that Tyr-180 is a catalytic residue, Lys-97 and His-79 function both coenzyme binding and chemical steps of the reaction, and Tyr-71 and His-76 are involved in coenzyme binding. In addition, Trp-125, Asp-176 and Trp-254 were suggested to be responsible for substrate binding, and of the three residues Asp-176 is thought to be important for the adaptation of the alcohol substrate in to the binding site. The crystallographic study is now in progress.4. Dimeric DD was inhibited by several drugs such as nonsteroidal anti-inflammatory agents, of which the potent inhibitors commonly had 4-hydroxyphenylketone structure. The enzyme's mRNA, although it did not detected in normal human kidney, was detected in one of five renal cancer specimen. The variant gene with respect to exon 4 region was not detected, and such a study for other exon regions will be continued to find clinical significance.5. During the course of cloning of the genomic gene for dimeric, I noticed that the gene is included in a working draft sequence of chromosome 19. The 5'-noncoding region of this gene contains various putative binding sites for transcription factors.6. The structural and functional aspects obtained in this study was presented in 10^<th> International Symposium on Enzymology and Molecular Biology of Carbonyl Metabolism (New Mixico, U.S.A., July, 2000). Less

二氢二醇脱氢酶(DD，EC 1.3.1.20)催化依赖于NADP氧化芳香烃的反式二氢二醇，参与了芳香烃的细胞毒性。这种酶以多种形式存在于哺乳动物组织中。在多种形式中，二聚体DD是组织特异性表达的，但其结构和生理功能尚未阐明。本研究旨在阐明二聚体DD的结构及其与功能的关系和可能的临床意义。对所获得的结果进行了总结。猴肾、猪和狗肝中的二聚体DD与依赖NADP的D-木糖脱氢酶(EC1.1.1.179)相同。对二聚体DDs在兔晶状体、人肠道和上述动物组织中的蛋白质测序和/或克隆的一级结构表明，这些酶与包括葡萄糖-果糖氧化还原酶在内的原核蛋白组成了一个新的蛋白质家族。The Site…对猴二聚体DD的更多定向突变表明，Tyr-180是一个催化残基，Lys-97和His-79既具有辅酶结合功能，又具有反应的化学步骤，Tyr-71和His-76参与辅酶结合。此外，Trp-125、Asp-176和Trp-254被认为负责底物结合，其中Asp-176被认为对乙醇底物与结合部位的适应起重要作用。结晶学研究目前正在进行中。二聚体DD可被非甾体抗炎药等多种药物抑制，其中有效的抑制药一般具有4-羟基苯基酮结构。尽管在正常人肾脏中没有检测到这种酶的mRNA，但在五例肾癌标本中有一例检测到了这种酶的mRNA。未检测到与外显子4区域相关的变异基因，对其他外显子区域的研究将继续具有临床意义。在克隆二聚体基因组基因的过程中，我注意到该基因包含在19号染色体的工作草案序列中，该基因的5‘-非编码区含有各种可能的转录因子结合位点。本研究获得的结构和功能方面已在第10届国际羰基代谢的酶学和分子生物学研讨会(美国New Mixico，2000年7月)上发表。较少

项目成果

E.Arimitsu et al.: "Cloning and sequencing of the cDNA species for mammalian dihydrodiol dehydrogenases"Biochem.J.. 342・3. 721-728 (1999)

E.Arimitsu等：“哺乳动物二氢二醇脱氢酶的cDNA种类的克隆和测序”Biochem.J. 342·3(1999)。

S.Aoki et al.: "Identity of dimeric dihydrodiol dehydrogenase as NADP^+-dependent D-xylose dehydrogenase in pig liver"Chemico-Biol.Interact.. (in the press). (2001)

S.Aoki等人：“猪肝中二聚二氢二醇脱氢酶作为NADP 2 -依赖性D-木糖脱氢酶的身份”Chemico-Biol.Interact..（出版中）。

Y.Asada et al.: "Roles of His-79 and Tyr-180 of D-xylose/dihydrodiol Dehydrogenase in catalytic function"Biochem.Biophys.Res.Commun.. 278-2. 333-337 (2000)

Y.Asada 等：“D-木糖/二氢二醇脱氢酶的 His-79 和 Tyr-180 在催化功能中的作用”Biochem.Biophys.Res.Commun. 278-2。

S.Aoki et al.: "Identity of dimeric dihydrodiol dehydrogenase as NADP^+-dependent D-xylose dehydrogenase in pig liver"Chemico-Biol.Interact.. (印刷中). (2001)

S. Aoki 等人：“猪肝中二聚二氢二醇脱氢酶作为 NADP + 依赖性 D-木糖脱氢酶的身份”Chemico-Biol.Interact..（印刷中）。