NUTRITIONAL REGULATION OF SELENOENZYMES AND PROTEINS

本申请建议研究硒的营养调节-- 含有酶和蛋白质的。谷胱甘肽过氧化物酶(GPX)是最好的 动物体内存在的特有的硒酶，因此这项研究将 主要关注GPX。GPX活动水平在以下期间迅速下降 饮食中的硒缺乏，我们已经使用了抗GPX抗体来证明 GPX蛋白水平也呈指数级协调下降 进行性缺硒。用克隆的GPX基因在Northern印迹中证明 分析，我们最近表明，饮食中的硒水平调节 GPx的mRNA水平以及GPx的活性和蛋白，因此表明 动物的硒状态可能调节含硒的酶，如GPX在 基因表达的转录水平。这一监管机制 也许这是为什么大多数人对膳食中的硒的需求 通常使用GPX作为指标来确定的高等动物也是如此 很相似。 在过去的十年里，越来越明显的是，一些 除GPX外，硒蛋白还存在于高等动物组织中 但这些硒蛋白的性质和功能在很大程度上是 未知。与其他依赖微量饮食因素的物种一样 为了充分发挥生物活性，这些物种应该进一步 特色化的。因为活性分析、抗体或cdna探针不是 普遍适用于这些未鉴定的硒蛋白，我们的 先前开发的SDS/PAGE程序可用于识别和 对这些硒蛋白中的一些进行75Se标记。这将是 允许对这些相对没有特征性的硒蛋白进行初步研究。 为了加深我们对硒代谢的理解，我们打算继续 以下是具体目标：1)确定硒状况对 GPX(信使核糖核酸、蛋白质和酶活性水平)的表达 真核生物，并进一步表征这一过程；2)提纯和 进一步表征一个或多个未表征的哺乳动物 硒蛋白。 这项研究的总体目标是更好地理解 硒蛋白和硒酶如GPX的营养调节。 这一知识可能会使我们更好地理解 硒的生物化学及其对富硒的认识 在动物和人类健康中的作用。

This application proposes to study the nutritional regulation of selenium- containing enzymes and proteins. Glutathione peroxidase (GPX) is the best characterized selenoenzyme present in animals, and thus this research will have a major focus on GPX. GPX activity levels decrease rapidly during dietary Se deficiency, and we have used anti-GPX antibodies to show that GPX protein levels also fall exponentially and coordinately during progressive Se deficiency. Using a cloned GPX cDNA prove in Northern blot analysis, we have recently shown that the level of Se in the diet regulates the level of GPX mRNA as well as GPX activity and protein, thus suggesting that Se status of animals may regulate Se-containing enzymes such as GPX at the transcriptional level of gene expression. This regulatory mechanism maybe one of the major reasons why the dietary Se requirements for most higher animals, usually determined using GPX as an indicator, are so similar. In the last decade it has become increasingly clear that a number of selenoproteins in addition to GPX are present in the tissues of higher animals, but the nature and function of these selenoproteins are largely unknown. As with other species that are dependent on trace dietary factors for full biological activity, these species should be further characterized. Because activity assays, antibodies or cDNA probes are not generally available for these uncharacterized selenoproteins, our previously developed SDS/PAGE procedure can be used to identify and to quantitate 75Se-labeling of a number of these selenoproteins. This will allow initial studies on these relatively uncharacterized selenoproteins. To further our understanding of Se metabolism, we intend to pursue the following specific objectives: 1) to determine the effect of Se status on the expression of GPX (mRNA, protein and enzyme activity levels) in eukaryotes, and to further characterize this process; 2) to purify and to further characterize one or more of the uncharacterized mammalian selenoproteins. The overall objective of this research is to better understand the nutritional regulation of selenoproteins and selenoenzymes such as GPX. This knowledge may lead to a better understanding of the nutritional biochemistry of Se and it may increase our understanding of selenium's full role in animal and human health.