2型猪链球菌表面蛋白 Hp0272的原核表达及其结合人血清 IgA 的活性分析

目的：构建带GST标签的2型猪链球菌功能未知表面蛋白Hp0272的原核表达质粒并在大肠杆菌中表达，获得纯度较高的GST-Hp0272重组融合蛋白，鉴定其与人血清IgA（higA）的结合活性。方法：采用分子生物学方法构建pGEX-4T-1-0272重组表达载体，将其转入大肠杆菌BL21（DE3）菌株，IPTG诱导表达后，通过SDS-PAGE分析蛋白表达情况；用GSTrap柱亲和纯化目的蛋白并经Western印迹验证，采用配体印迹和ELISA方法鉴定GST-Hp0272与hlgA的结合活性。结果：构建了GST融合蛋白原核表达质粒，并获得GST-Hp0272融合蛋白，鉴定到Hp0272特异性地与hisA结合，且结合区域位于Hp0272的N端（41-318an）。结论：获得了GST-Hp0272融合蛋白，并鉴定到其能特异性结合hlgA，为进一步了解Hp0272在猪链球菌致病过程中的作用提供了基础。

Prokaryotic Expression and hIgA-Binding Activity Identification of Recombinant Streptococcus suis Serotype 2 Surface Protein Hp0272

Objective： To construct prokaryotic expression vector, express and purify the GST-tag fusion protein Hp0272（a function unknown suface protein） of Streptococcus suis serotype 2 and identify its binding activity with human serum IgA（hIgA）. Methods： The gene encoding Hp0272 was cloned in plasmid pGEX-4T-1 and trans- formed into Escherichia coli BL21 to express fusion protein GST-Hp0272. The expression of fusion protein was an- alyzed by SDS-PAGE, frequently purified by GSTrap and identified by Western-blotting. In addition, the specific binding activities with hIgA was identified by Ligand-blotting and ELISA. Results： The prokaryotic expression vec- tor of GST-Hp0272 was constructed. The pure fusion protein GST-Hp0272 was expressed and purified by affinity purification. The specific binding activity with hIgA was identified and the domain located on the N-Terminal （41- 318 aa）. Conclusion： GST-Hp0272 was obtained by prokaryotic expression and its binding activity with hIgA was identified, which will help us to futher study the roles of Hp0272 in pathogenesis of S.suis.