Entamoeba histolytica EhDEAD1 is a conserved DEAD-box RNA helicase with ATPase and ATP-dependent RNA unwinding activities
Entamoeba histolytica EhDEAD1 is a conserved DEAD-box RNA helicase with ATPase and ATP-dependent RNA unwinding activities
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DOI：
10.1016/j.gene.2008.01.024
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发表时间：
2008-05-15
期刊：
影响因子：
3.5
通讯作者：
Orozco, Esther
中科院分区：
文献类型：
--
作者：
Lopez-Camarillo, Cesar;Garcia-Hernandez, Maria de la Luz;Orozco, Esther
RNA 解旋酶是广泛保守的关键酶，在 RNA 代谢中发挥多种功能。在这里，我们介绍了原生动物寄生虫溶组织内阿米巴中 EhDEAD1 RNA 解旋酶的克隆、表达和功能特征。根据其一级结构，EhDEAD1 与酵母 DED1 和人 DDX3X RNA 解旋酶在进化上相关，两者都参与翻译和细胞周期调节。 EhDEAD1 预测的氨基酸序列显示了其他生物体中报道的 DEAD-box SFII 超家族成员的九个保守基序，并且它在进化上与原生动物同源物接近。纯化的重组 EhDEAD1 蛋白具有 ATPase 活性，并且能够在体外以 ATPase 依赖性方式结合和解旋 RNA。 RT-PCR检测表明EhDead1基因在细胞周期S期过度转录。此外，反义RNA抑制EhDead1基因表达似乎促进了从S期向G2/M期的转变。有趣的是，我们的结果表明，尽管与酵母 DED1 具有高度的序列同源性，但 EhDEAD1 无法拯救翻译中受影响的两种酵母 Ded1 RNA 解旋酶突变体。 (c) 2008 Elsevier B.V. 保留所有权利。
RNA helicases are widely conserved key enzymes that perform multiple functions in RNA metabolism. Here, we present the cloning, expression and functional characterization of the EhDEAD1 RNA helicase in the protozoan parasite Entamoeba histolytica. According to its primary structure, EhDEAD1 is evolutionary related to yeast DED1 and human DDX3X RNA helicases, both involved in translation and cell cycle regulation. The EhDEAD1 predicted amino acid sequence exhibits the nine conserved motifs described for the DEAD-box SFII superfamily members reported in other organisms and it is evolutionary close to protozoan homologues. Purified recombinant EhDEAD1 protein presented ATPase activity and it was able to bind and unwind RNA in an ATPase-dependent manner in vitro. RT-PCR assays showed that EhDead1 gene is overtranscribed in the cell cycle S phase. Moreover, inhibition of EhDead1 gene expression by antisense RNA seemed to facilitate transition from S to G2/M phase. Intriguingly, our results showed that EhDEAD1 was unable to rescue two yeast Ded1 RNA helicase mutants affected in translation, in spite of the high sequence homology with yeast DED1. (c) 2008 Elsevier B.V. All rights reserved.