Biosynthesis of Selenocysteine and Its Incorporation into Protein

硒是人类和其他哺乳动物饮食中必需的微量营养素， 有许多健康的好处已被归因于这种元素，包括预防癌症，心脏病， 疾病和其他心血管和肌肉疾病，抑制病毒表达，延缓 艾滋病在艾滋病毒阳性患者中的发展，减缓衰老过程， 在哺乳动物发育、雄性生殖和免疫功能中的作用。我们先前提出， 硒的健康益处在很大程度上是由于 硒蛋白是含硒氨基酸，硒代半胱氨酸（Sec）。因为利特尔 我们知道Sec是如何生物合成的，我们进行了一个项目来阐明这种氨基是如何合成的。 氨基酸，这是遗传密码中的第21个氨基酸，被合成并鉴定， 表征参与该途径的每个组分。我们建立了生物合成 真核生物和古菌中Sec的途径（PLoS Biology 5：96-105，2007），并继续我们的研究。 负责Sec合成的组分的表征。在过去的一年里，我们 已经确定参与Sec生物合成途径的酶可用于 使用硫化物作为底物在tRNA[Ser]Sec上合成Cys。Cys-tRNA[Ser]Sec可以是 用于在硒蛋白翻译中解码码字UGA时插入Cys。该途径 在硒的饮食条件下， 哺乳动物细胞中的缺陷。我们还产生了两种条件性敲除小鼠模型， 硒磷酸合成酶1（SPS 1）和SECp 43敲除小鼠。两条鼠标线都表明， 相应基因的完全敲除是胚胎致死的。我们发现SPS 1 肝敲除不影响硒蛋白代谢，而SECp 43肝敲除影响硒蛋白代谢。 仍在调查中我们正在对SPS 1基因敲除小鼠进行胚胎分析， SPS 1在哺乳动物胚胎发育中的作用我们还在哺乳动物中发现了一种新的因子， 可能参与tRNA[Ser]Sec修饰的细胞，这种修饰可以 显著增加丝氨酸与tRNA[Ser]Sec的丝氨酰-tRNA合成酶连接。活动 在酵母丝氨酰-tRNA合成酶中没有发现这种因子，有趣的是， 酵母不编码硒代半胱氨酸生物合成机制。我们正在提纯这个因子， 表征其在修饰tRNA[Ser]Sec中的作用。

Selenium is an essential micronutrient in the diet of humans and other mammals and it has many health benefits have been ascribed to this element including preventing cancer, heart disease and other cardiovascular and muscle disorders, inhibiting viral expression, delaying the progression of AIDS in HIV positive patients, slowing the aging process, and having roles in mammalian development, male reproduction and immune function. We proposed previously that the health benefits of selenium are due in large part to the presence of selenium in selenoproteins as the selenium-containing amino acid, selenocysteine (Sec). Since little was known about how Sec was biosynthesized, we undertook a project to elucidate how this amino acid, which is the 21st amino acid in the genetic code, was synthesized and to identify and characterize each of the components involved in the pathway. We established the biosynthetic pathway of Sec in eukaryotes and archeae (PLoS Biology 5: 96-105, 2007) and are continuing our characterization of the components responsible for Sec synthesis. During the past year, we have identified that the enzymes involved in the biosynthetic pathway of Sec can be used to synthesize Cys on the tRNA[Ser]Sec using sulfide as a substrate. The Cys-tRNA[Ser]Sec can be used to insert Cys upon decoding the codeword, UGA, in selenoprotein translation. This pathway of misincorporating Cys into protein is likely used under dietary conditions of selenium deficiency in mammalian cells. We have also generated two conditional knock-out mouse models, selenophosphate synthetase 1 (SPS1) and SECp43 knockout mice. Both mouse lines have shown that the total knockout of the corresponding gene is embryonic lethal. We have found that the SPS1 liver knockout did not affect selenoprotein metabolism while the SECp43 liver knockout is still being investigated. We are conducting embryo analysis of SPS1 knockout mice to elucidate the function of SPS1 in mammalian embryo development. We also found a new factor in mammalian cells which may be involved in tRNA[Ser]Sec modification, and this modification can dramatically increase seryl-tRNA synthetase attachment of serine to tRNA[Ser]Sec. The activity of this factor was not found to be present in yeast seryl-tRNA synthetase, and interestingly, yeast do not encode the selenocysteine biosynthesis machinery. We are purifying this factor to characterize its role in modifying tRNA[Ser]Sec.