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首页> 外文学位 >A physiological, biochemical and structural analysis of inositol polyphosphate 5-phosphatases from Arabidopsis thaliana and humans.
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A physiological, biochemical and structural analysis of inositol polyphosphate 5-phosphatases from Arabidopsis thaliana and humans.

机译：拟南芥和人类肌醇多磷酸5-磷酸酶的生理，生化和结构分析。

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摘要

The complete role of inositol signaling in plants and humans is still elusive. The plant Arabidopsis thaliana contains fifteen predicted inositol polyphosphate 5-phosphatases (5PTases, E.C. 3.1.3.36) that have the potential to remove a 5-phosphate from various inositol second messenger substrates. To examine the substrate specificity of one of these Arabidopsis thaliana 5PTases (At5PTases), recombinant At5PTase1 was obtained from a Drosophila melanogaster expression system and analyzed biochemically. This analysis revealed that At5PTase1 has the ability to catalyze the hydrolysis of four potential inositol second messenger substrates.;To determine whether At5PTase1 can be used to alter the signal transduction pathway of the major drought-sensing hormone abscisic acid (ABA), plants ectopically expressing At5PTase1 under the control of a constitutive promoter were characterized. This characterization revealed that plants ectopically expressing At5PTase1 had an altered response to ABA. These plants have stomata that are insensitive to ABA, and have lower basal and ABA-induced inositol (1,4,5)-trisphosphate [Ins(1,4,5)P3] levels. In addition, At5PTase1 mRNA and protein levels are transiently regulated by ABA. These data strongly suggest that At5PTase1 can act as a signal terminator of ABA signal transduction.;Like the Arabidopsis At5PTase1, a human 5PTase, Ocrl, has the ability to catalyze the hydrolysis of a 5-phosphate from several inositol-containing substrates. The loss of functional Ocrl protein results in a rare genetic disorder known as Lowe oculocerebrorenal syndrome. To gather information concerning the specificity determinants of the Ocrl protein, a structure-function analysis of Ocrl was conducted using a vibrational technique, difference Fourier transform infrared (FT-IR) spectroscopy. Upon the introduction of Ins(1,4,5)P3 substrate, structural changes in carboxylic acid and histidine residues were observed. The net result of changes in these residues indicates that upon Ins(1,4,5)P3 introduction, a carboxylic acid-containing residue is protonated, and a histidine residue is deprotonated. This interpretation supports the idea that the deprotonation of the histidine residue is concomitant with the coordination of a divalent cation upon Ins(1,4,5)P3 introduction. This work allows for the proposal of a new model for the role of the active site histidine of OCRL.
机译：肌醇信号在植物和人类中的完整作用仍然难以捉摸。植物拟南芥含有十五种预测的肌醇多磷酸5-磷酸酶（5PTase，E.C。3.1.3.36），具有从各种肌醇第二信使底物中去除5-磷酸的潜力。为了检查这些拟南芥5PTase（At5PTases）之一的底物特异性，从果蝇表达系统获得了重组At5PTase1并进行了生化分析。该分析表明At5PTase1具有催化四种潜在的肌醇第二信使底物水解的能力;确定At5PTase1是否可用于改变异位表达植物的主要干旱敏感激素脱落酸（ABA）的信号转导途径。在组成型启动子的控制下At5PTase1进行了表征。此特征表明异位表达At5PTase1的植物对ABA的反应发生了变化。这些植物的气孔对ABA不敏感，并且具有较低的基础和ABA诱导的肌醇（1,4,5）-三磷酸[Ins（1,4,5）P3]水平。另外，At5PTase1 mRNA和蛋白质水平受ABA瞬时调节。这些数据强烈暗示At5PTase1可以充当ABA信号转导的信号终止子。与拟南芥At5PTase1一样，人类5PTase Ocrl具有催化从几种含肌醇的底物中水解5-磷酸的能力。功能性Ocrl蛋白的丧失会导致一种罕见的遗传疾病，称为Lowe眼脑肾综合征。为了收集有关Ocr1蛋白特异性决定子的信息，使用振动技术，差分傅里叶变换红外（FT-IR）光谱进行了Ocr1的结构功能分析。引入Ins（1,4,5）P3底物后，观察到羧酸和组氨酸残基的结构变化。这些残基变化的最终结果表明，引入Ins（1,4,5）P3后，含羧酸的残基被质子化，而组氨酸残基被去质子化。这种解释支持这样的想法，即在Ins（1,4,5）P3引入后，组氨酸残基的去质子化与二价阳离子的配位相伴。这项工作允许为OCRL的活性位点组氨酸的作用提出一个新模型的建议。

著录项

  • 作者

    Burnette, Ryan N.;

  • 作者单位

    Virginia Polytechnic Institute and State University.;

  • 授予单位 Virginia Polytechnic Institute and State University.;
  • 学科 Biology Molecular.;Biophysics General.;Chemistry Biochemistry.
  • 学位 Ph.D.
  • 年度 2004
  • 页码 170 p.
  • 总页数 170
  • 原文格式 PDF
  • 正文语种 eng
  • 中图分类
  • 关键词

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