Pham X H, Reddy M K, Ehtesham N Z, Matta B, Tuteja N
Plant Molecular Biology Group, International Centre for Genetic Engineering and Biotechnology, PO Box 10504, Aruna Asaf Ali Marg, New Delhi 110 067, India.
Plant J. 2000 Oct;24(2):219-29. doi: 10.1046/j.1365-313x.2000.00869.x.
DNA helicases play an essential role in all aspects of nucleic acid metabolism, by providing a duplex-unwinding function. This is the first report of the isolation of a cDNA (1.6 kb) clone encoding functional DNA helicase from a plant (pea, Pisum sativum). The deduced amino-acid sequence has eight conserved helicase motifs of the DEAD-box protein family. It is a unique member of this family, containing DESD and SRT motifs instead of DEAD/H and SAT. The encoded 45.5 kDa protein has been overexpressed in bacteria and purified to homogeneity. The purified protein contains ATP-dependent DNA and RNA helicase, DNA-dependent ATPase, and ATP-binding activities. The protein sequence contains striking homology with eIF-4A, which has not so far been reported as DNA helicase. The antibodies against pea helicase inhibit in vitro translation. The gene is expressed as 1.6 kb mRNA in different organs of pea. The enzyme is localized in the nucleus and cytosol, and unwinds DNA in the 3' to 5' direction. The pea helicase interacts with pea topoisomerase I protein and stimulates its activity. These results suggest that pea DNA helicase could be an important multifunctional protein involved in protein synthesis, maintaining the basic activities of the cell, and in upregulation of topoisomerase I activity. The discovery of such a protein with intrinsic multiple activity should make an important contribution to our better understanding of DNA and RNA transactions in plants.
DNA解旋酶通过提供双链解旋功能，在核酸代谢的各个方面发挥着至关重要的作用。这是首次从植物（豌豆， Pisum sativum）中分离出编码功能性DNA解旋酶的cDNA（1.6 kb）克隆的报告。推导的氨基酸序列具有DEAD-box蛋白家族的八个保守解旋酶基序。它是该家族的一个独特成员，包含DESD和SRT基序而非DEAD/H和SAT。编码的45.5 kDa蛋白已在细菌中过表达并纯化至同质。纯化的蛋白含有ATP依赖性DNA和RNA解旋酶、DNA依赖性ATP酶以及ATP结合活性。该蛋白序列与eIF-4A具有显著同源性，而eIF-4A迄今尚未被报道为DNA解旋酶。针对豌豆解旋酶的抗体抑制体外翻译。该基因在豌豆的不同器官中表达为1.6 kb mRNA。该酶定位于细胞核和细胞质中，并沿3'至5'方向解旋DNA。豌豆解旋酶与豌豆拓扑异构酶I蛋白相互作用并刺激其活性。这些结果表明，豌豆DNA解旋酶可能是一种重要的多功能蛋白，参与蛋白质合成、维持细胞的基本活性以及上调拓扑异构酶I的活性。发现这样一种具有内在多种活性的蛋白，应为我们更好地理解植物中的DNA和RNA交易做出重要贡献。
