CAJ | 학술논문

为了获得PRRSV非结构蛋白2-半胱氨酸结构域的高纯度原核表达蛋白以及了解该蛋白的抗原表位，试验以从PRRSV-VR2332毒株上提取的病毒全基因组RNA为模板，通过RT-PCR扩增非结构蛋白2-半胱氨酸结构域（Nsp2pro）基因，连接构建原核表达载体SUMO-Nsp2pro，转入宿主茵Rosetta2，经IPTG诱导，获得高浓度的可溶蛋白Nsp2pro，经亲和层析His-Bind后得到高纯度的目的蛋白。同时，运用生物信息学方法对Nsp2pro二级结构及抗原表位进行预测。结果显示，Nsp2pro的α螺旋及β折叠区域较多，转角区域较少，结构较为复杂，位于蛋白分子表面且亲水的区段很可能是B细胞表位的优势区段。获得较高纯度的蛋白，将为后续进一步研究Nsp2pro基因编码的蛋白在病毒复制过程中的作用奠定基础。
위료획득PRRSV비결구단백2-반광안산결구역적고순도원핵표체단백이급료해해단백적항원표위，시험이종PRRSV-VR2332독주상제취적병독전기인조RNA위모판，통과RT-PCR확증비결구단백2-반광안산결구역（Nsp2pro）기인，련접구건원핵표체재체SUMO-Nsp2pro，전입숙주인Rosetta2，경IPTG유도，획득고농도적가용단백Nsp2pro，경친화층석His-Bind후득도고순도적목적단백。동시，운용생물신식학방법대Nsp2pro이급결구급항원표위진행예측。결과현시，Nsp2pro적α라선급β절첩구역교다，전각구역교소，결구교위복잡，위우단백분자표면차친수적구단흔가능시B세포표위적우세구단。획득교고순도적단백，장위후속진일보연구Nsp2pro기인편마적단백재병독복제과정중적작용전정기출。
In order to acquire highly purified protein of cysteine domain of nonstructural protein 2 of PRRSV and acquaintance its antigen epiopes, the experiment were conducted with the genome extracted from PRRSV-VR2332 and got the gene of the cysteine domain of nonstructural protein 2 of PRRSV（Nsp2pro）by RT-PCR. Then,a recombinant plasmid named SUMO-Nsp2pro had been constructed, and then transformed SUMO-Nsp2pro into Rosetta2. Therefore,a large amount of Nsp2pro was obtained by IPTG. In the end,Nsp2pro was purified by His-Bind affinity chromatography,meanwhile predicted the second structure and antigen epiopes through means of hio-information,the results showed that Nsp2pro owned a great many Alpha regions and Beta regions while few of turn regions. The antigen epitopes were located in hydrolicity regions possibly. The obtained high purified protein would provide foundations for the further researches on Nsp2pro gene.