HBcAg和HBsAg前S1表位肽融合蛋白的表达研究

构建、表达并纯化了HBcAg与HBsAg前S1表位肽融合蛋白BTcs1，为开发新型HBV治疗和预防性疫苗提供实验依据。利用DNA重组技术，构建了HBcAg与HBsAg前S1表位肽融合蛋白原核表达质粒pBTcs1，在大肠杆菌（HB101）中进行表达，用蔗糖密度梯度超速离心对表达产物BTcs1进行纯化，用SDS-PAGE、SEC、WESTERN-BLOT和电镜进行鉴定。结果表明成功构建了HBcAg与HBsAg前S1表位肽融合蛋白原核表达质粒pBTcs1，BTcs1的表达量为20-25 mg/L，DOT-BLOT结果显示BTcs1主要分布在30%-50%蔗糖介质中，SDS-PAGE和SEC结果显示蛋白纯度>95%，WESTERN-BLOT结果显示BTcs1可以与抗HBcAg抗体和抗HBsAg前S1抗体特异杂交，显色条带在约28 kD处，电镜分析表明BTcs1可以自主组装成病毒样颗粒（VLP），直径约为30-34 nm。本研究为进一步探讨BTcs1的功能及应用奠定了基础。

The study of expression HBcAg and HBsAg preS1 epitope peptide fusion protein

To construct, express and purify fusion protein containing HBcAg and HBsAg preS1 epitope peptide for the purpose of investigating a novel HBV vaccine with both prophylactic and therapeutic functions. Using DNA recombinant technology, prokaryotic expression plasmid pBTcs1 expressing HBcAg and HBsAg pre-S1 epitope peptide fusion protein was constructed. After expressed in E.coli. HB101, the production BTcs1 was purified by sucrose density gradient ultracentrifugation and identified by SDS-PAGE, SEC, Western-blot and electron microscope. The results indicated that expression plasmid pBTcs1 was constructed successfully, and 20~25 mg purified BTcs1 fusion protein was obtained from 1L LB culture. Result of DOT-BLOT indicated that the distribution of BTcs1 was mainly in 30~50% sucrose, the purity of BTcs1 was greater than 95% by SDS-PAGE and SEC analysis. BTcs1 could probe with specific antibodies at 28 kDa by Western-blot, BTcs1 could also self assemble VLP by electron microscope analysis, its diameter was 30~34 nm approximately. The present study lay a foundation for further research functions and applications of BTcs1.