Dynamics of the essential trace element selenium in mammals and molecular basis for selenoprotein biosynthesis

哺乳动物必需微量元素硒的动态及硒蛋白生物合成的分子基础

基本信息

批准号：
17370037
负责人：
ESAKI Nobuyoshi
金额：
$ 9.47万
依托单位：
Kyoto University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2005
资助国家：
日本
起止时间：
2005 至 2006
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-17370037/
关键词：
selenium selenoprotein selenocysteine essential trace element enzyme 翻訳 SECIS セレノシステインリアーゼ

项目摘要

Selenium is an essential trace element with a variety of physiological role and is strictly discriminated from its cognate sulfur to be incorporated into a specific position of proteins as a selenocysteine residue. Mammalian cells are proposed to contain 25 selenoproteins. Selenocysteine residues are not directly synthesized from free selenocysteine but are biosynthesized from an active form of selenium which is formed from selenocysteine by decomposition at the translation step. In the current study, we identified factors involved in selenoprotein biosynthesis in mammalian cells and analyzed their functions.1. We identified nucleolin and RAD17 as novel proteins which sqiecifically recognize and bind to selenocysteine insertion sequence (SECTS).2. Selenccystiene lyase (SCL) decomposes L-selenocysteine to form selenium and L-alanine and is involved in selenoprotein biosynthesis. We salved X-ray crystal structure of SCL from rat in the native form and in the complexed form with L-cysteine. In the structure of SCL complexed with L-cysteine, the thiol group of L-cysteine was coordinated to bind to PLP-lysine Shiff base of the enzyme. The result suggest that Cys 375 of SCL plays an important role in the strict substrate specificity of the enzyme toward L-selenocysteine.3. We constructed the mutants of selenophosphate synthetase (SeID) from Escherichia coli by site directed mutagenesis. The analysis showed that a highly conserved cysteine residue (Cys17) of SeID is essential for the catalytic activity of the enzyme.

硒是一种具有多种生理作用的必需微量元素，与其同源硫严格区别，以硒代半胱氨酸残基的形式掺入蛋白质的特定位置。哺乳动物细胞被认为含有 25 种硒蛋白。硒代半胱氨酸残基不是直接由游离的硒代半胱氨酸合成的，而是由活性形式的硒生物合成的，硒是由硒代半胱氨酸在翻译步骤中分解形成的。在本研究中，我们鉴定了哺乳动物细胞中参与硒蛋白生物合成的因子并分析了它们的功能。 1．我们将核仁素和RAD17鉴定为新型蛋白质，它们专门识别并结合硒代半胱氨酸插入序列(SECTS)。2．硒半胱氨酸裂解酶 (SCL) 分解 L-硒代半胱氨酸形成硒和 L-丙氨酸，并参与硒蛋白的生物合成。我们保存了大鼠 SCL 的 X 射线晶体结构，包括天然形式和与 L-半胱氨酸的复合形式。在与L-半胱氨酸复合的SCL结构中，L-半胱氨酸的硫醇基团与酶的PLP-赖氨酸希夫碱基配位结合。结果表明，SCL的Cys 375在该酶对L-硒代半胱氨酸的严格底物特异性中起重要作用。 3．我们通过定点诱变从大肠杆菌中构建了硒磷酸合成酶（SeID）突变体。分析表明，SeID 的高度保守的半胱氨酸残基（Cys17）对于酶的催化活性至关重要。