Department of Biotechnology, Anna University, BIT Campus, Tiruchirappalli 620 024, India.
Biomed Res Int. 2013;2013:343195. doi: 10.1155/2013/343195. Epub 2013 Sep 8.
Detection of nonstructural protein (NS1) is an important diagnostic marker during acute phase of dengue infection. Not only for diagnostic purpose, the protein had important role in vaccine design as well, as a candidate for studying virus assembly and maturation. Various researchers employed different expression systems and strategies for recombinant NS1 protein production. Attempts to express NS1 protein in prokaryotic and yeast expression system result in formation of insoluble protein which needs to undergo refolding to attain native structural and functional forms. Here, we report the production of soluble NS1 protein in E. coli by using appropriate vector and employing suitable culture conditions to maximize protein production. Proteins were purified using metal affinity chromatography. SDS-PAGE and western blot analysis reveal the native structure of NS1 protein. Solid phase ELISA using the recombinantly expressed antigen with positive and negative dengue samples showed that the expressed protein retains its antigenic and immunological properties. To our knowledge, this is the first report on the successful production of functionally active recombinant dengue-2 NS1 protein production without undergoing any in vitro posttranslational modification process.
非结构蛋白（NS1）的检测是登革热感染急性期的一个重要诊断标志物。该蛋白不仅可用于诊断目的，在疫苗设计中也具有重要作用，可作为研究病毒组装和成熟的候选物。许多研究人员采用不同的表达系统和策略来生产重组 NS1 蛋白。在原核和酵母表达系统中尝试表达 NS1 蛋白会导致形成不溶性蛋白，需要进行复性以获得天然的结构和功能形式。在这里，我们通过使用适当的载体和合适的培养条件在大肠杆菌中报告可溶性 NS1 蛋白的生产，以最大限度地提高蛋白产量。使用金属亲和层析对蛋白质进行纯化。SDS-PAGE 和 Western blot 分析显示 NS1 蛋白的天然结构。使用与阳性和阴性登革热样本的重组表达抗原进行固相 ELISA 表明，表达的蛋白保留其抗原性和免疫原性。据我们所知，这是首次成功生产无任何体外翻译后修饰过程的功能性重组登革热 2 型 NS1 蛋白的报告。
