• 主题
高级搜索  >
历史搜索 清空历史
首页> 外文学位 >The role of serine acetyltransferase in nickel and selenium assimilation and tolerance in metal hyperaccumulators.
【24h】

The role of serine acetyltransferase in nickel and selenium assimilation and tolerance in metal hyperaccumulators.

机译：丝氨酸乙酰转移酶在镍和硒同化中的作用以及金属超富集剂的耐受性。

获取原文
获取原文并翻译 | 示例
页面导航
  • 摘要
  • 著录项
  • 相似文献
  • 相关主题

摘要

When growing in its native habitat, hyperaccumulators can accumulate two to three orders of magnitude higher concentration of metals in their shoots when compared to other plants grown in the same habitat, without showing symptoms of metal toxicity.;We previously reported that the constitutively elevated concentration of the antioxidant glutathione (GSH) is involved in the ability of Ni hyperaccumulating Thlaspi species to protect against nickel (Ni)-induced oxidative stress. Given the central role of SAT in regulating sulfur (S) assimilation and GSH biosynthesis we propose that SAT is differentially regulated in Thlaspi goesingense to allow for enhanced GSH biosynthesis. Specifically, we hypothesize that SAT in T. goesingense is less sensitive to inhibition by cysteine. Such reduced sensitivity to cysteine would provide a mechanism to allow enhanced steady-state levels of O-acetylserine (OAS), leading to increased cysteine and ultimately GSH biosynthesis. We have previously cloned three SAT cDNAs from T. goesingense. Based on the predicted amino acid sequences of these TgSATs, and their alignment to the SATs from the nonaccumulator A. thaliana, we predicted which SAT isoform was homologous to the cysteine sensitive cytosolic SAT from A. thaliana. We found that recombinant cytosolic SAT from T. goesingense (TgSAT-c) is not inhibited by cysteine unlike the homologous cytosolic SAT from A. thaliana (AtSAT-c) which is sensitive to cysteine. Using domain swapping and site-directed mutagenesis we identified two amino acid residues in the C-terminus of TgSAT-c that are critical for the cysteine insensitivity of this enzyme compared to AtSAT-c. A change of cysteine to proline is most critical for this loss of cysteine sensitivity, with alanine to glycine playing a secondary role. Moreover, we also observed that this proline residue in TgSAT-c was also related to the competitive binding site for cysteine and the SAT substrate serine. Heterologous expression of the engineered and native cysteine insensitive SAT was observed to lead to increased Ni tolerance in both E. coli and A. thaliana when compared to expression of the cysteine sensitive SAT. Interestingly, the proline and alanine residues in SAT-c are well conserved in Thlaspi species, occurring in hyperaccumulator and nonaccumulator species, and this may partially explain the elevated OAS and GSH contents we observe across the Thlaspi genus. Furthermore, this may suggest why the Thlaspi genus appears to be evolutionally preadapt for Ni hyperaccumulation.;Selenium (Se) hyperaccumulators also have an elevated OAS content compared to nonaccumulators. Given the central role of OAS in S assimilation, we hypothesized that this elevated OAS plays an important role in up-regulating selenate uptake and reduction in Se hyperaccumulating species of Astragalus. Furthermore, we hypothesized that differential regulation of SAT in the Se hyperaccumulator leads to this elevation of OAS content compared to nonaccumulator. To directly test this, we cloned SAT cDNAs from the Se hyperaccumulator A. bisulcatus and the nonaccumulator A. drummondii. We found that recombinant cytosolic SAT from A. bisulcatus is less sensitive to cysteine and has a higher maximum enzyme activity compared to recombinant cytosolic SAT from A. drummondii. Furthermore, we found that the subcellular localization of SAT in A. bisulcatus differs from A. drummondii and this may also affect the production of OAS. Expression of Astragalus SAT in A. thaliana was not sufficient to produce Se hyperaccumulation. However, our preliminary results suggest that the expression of SAT and selenocysteine methyltransferase (SMT) together may be sufficient to induce Se hyperaccumulation. We demonstrate that the expression of SAT in combination with SMT leads to increased OAS, possibly up-regulating the expression of sulfate transporters, and increasing Se uptake in plants, although more work is needed to confirm these results.
机译：与在同一栖息地生长的其他植物相比，当在其原生栖息地中生长时，高累积量可在其芽中积累高2至3个数量级的金属，而不会表现出金属毒性的症状。谷胱甘肽（GSH）的抗氧化作用涉及Ni过度富集Thlaspi物种抵抗镍（Ni）诱导的氧化应激的能力。鉴于SAT在调节硫（S）同化和GSH生物合成中的核心作用，我们建议在Gosingense Goingense中对SAT进行差异调节，以增强GSH的生物合成。具体来说，我们假设戈恩根丝杉中的SAT对半胱氨酸的抑制作用较不敏感。对半胱氨酸的这种降低的敏感性将提供一种机制，以允许增加稳态水平的O-乙酰丝氨酸（OAS），从而导致半胱氨酸增加，并最终导致GSH的生物合成。我们之前已经从Goingense T.克隆了三个SAT cDNA。基于这些TgSATs的预测氨基酸序列，以及它们与非蓄积拟南芥中SAT的比对，我们预测哪种SAT亚型与拟南芥中半胱氨酸敏感的胞质SAT同源。我们发现，与来自拟南芥的同源胞质SAT（AtSAT-c）对半胱氨酸敏感的不同，来自戈氏菌的重组胞质SAT（TgSAT-c）不受半胱氨酸的抑制。使用域交换和定点诱变，我们在TgSAT-c的C末端鉴定了两个氨基酸残基，与AtSAT-c相比，这两个残基对该酶的半胱氨酸不敏感至关重要。对于这种半胱氨酸敏感性的丧失，将半胱氨酸变为脯氨酸是最关键的，丙氨酸向甘氨酸起次要作用。此外，我们还观察到，TgSAT-c中的脯氨酸残基也与半胱氨酸和SAT底物丝氨酸的竞争结合位点有关。与半胱氨酸敏感SAT的表达相比，工程和天然半胱氨酸不敏感SAT的异源表达可导致大肠杆菌和拟南芥中的Ni耐受性增加。有趣的是，SAT-c中的脯氨酸和丙氨酸残基在Thlaspi物种中非常保守，发生在超积累和非积累物种中，这可能部分解释了我们在Thlaspi属中观察到的OAS和GSH含量升高。此外，这可能暗示了为什么Thlaspi属在进化上似乎适合Ni过度富集。与非积累型硒相比，硒（Se）过度富集型的OAS含量也更高。鉴于OAS在S同化中的核心作用，我们假设这种升高的OAS在上调硒摄取和减少黄芪中Se富集物种中起重要作用。此外，我们假设与非蓄积器相比，硒超蓄积器中SAT的差异调节导致OAS含量升高。为了直接对此进行测试，我们从Se超级蓄积比目拟南芥和非蓄积的A. drummondii中克隆了SAT cDNA。我们发现，与来自A. drummondii的重组胞质SAT相比，来自A. bisulcatus的重组胞质SAT对半胱氨酸的敏感性较低，并且具有更高的最大酶活性。此外，我们发现SAT在比目拟南芥中的亚细胞定位不同于鼓膜拟南芥，这也可能影响OAS的产生。黄芪SAT在拟南芥中的表达不足以引起Se的过度积累。然而，我们的初步结果表明，SAT和硒代半胱氨酸甲基转移酶（SMT）一起表达可能足以诱导Se过度积累。我们证明，与SMT结合使用SAT的表达可增加OAS，可能上调硫酸盐转运蛋白的表达，并增加植物对Se的吸收，尽管需要更多的工作来确认这些结果。

著录项

  • 作者

    Na, GunNam.;

  • 作者单位

    Purdue University.;

  • 授予单位 Purdue University.;
  • 学科 Agriculture Horticulture.
  • 学位 Ph.D.
  • 年度 2009
  • 页码 175 p.
  • 总页数 175
  • 原文格式 PDF
  • 正文语种 eng
  • 中图分类
  • 关键词

相似文献

  • 外文文献
  • 中文文献
  • 专利
获取原文

客服邮箱：kefu@zhangqiaokeyan.com

京公网安备：11010802029741号 ICP备案号：京ICP备15016152号-6 六维联合信息科技 (北京) 有限公司©版权所有

  • 客服微信

  • 服务号