5个水稻蛋白激酶的克隆、表达及活性研究

蛋白激酶在生长发育、表达调控、逆境反应、信号传导等几乎所有的生命过程中发挥着重要作用,蛋白激酶的研究具有重要的理论和应用价值。全基因组测序结果表明,水稻基因组中至少有1500个蛋白激酶。本研究利用已有的蛋白激酶序列,通过PCR扩增和重组克隆,将5个重要的蛋白激酶在酿酒酵母系统中进行了表达,它们的编号分别是:NM-189878,NM-194133,NM-190508,NM-197522和NM-197571,其中NM-194133具有较强的自我磷酸化活性。同源性分析表明,这些激酶在水稻中可能的作用包括:碳代谢的生物合成调控、种子形态发生、细胞分裂调控、生物和非生物刺激的反应调控等。这项工作为进一步的体外酶活分析及蛋白质-蛋白质相互作用研究等提供了基础,也为高通量克隆表达水稻蛋白激酶提供了一个有效的系统。

Cloning, Expression and Autophosphorylation of 5 Rice Protein Kinases

It has been demonstrated that protein kinases play important roles in almost all biological process, e.g. growth and development, regulation of expression, stress responses and signal transduction. The finished rice genomic sequence indicated that at least 1500 protein kinases exist in rice genome. In the current study, five rice kinases has been amplified based their full-length cDNA sequences, their corresponding proteins have been expressed and purified in yeast, the accession number are NM-189878, NM-194133, NM-190508, NM-197522 and NM-197571 respectively. NM-194133 has strong autophosphorylation activity. Based on sequence similarity, the deduced functions of these kinases are biosynthesis regulation of carbon metabolism, seed embryogenesis, cell division regulation, response to biotic/abiotic stimulation etc. This work is helpful for in vitro enzyme assay, protein-protein interaction analysis, and provides an efficient system for high throughput cloning and expression of rice genes.