严重急性呼吸综合征冠状病毒S蛋白部分序列1的纯化及免疫活性分析

Purification of SARS Coronavirus spike protein fragment 1 and analysis of its immune activity

目的表达和纯化严重急性呼吸综合征(SARS)冠状病毒S蛋白部分序列1(S1),分析其诱导的免疫应答。方法大量诱导表达SARS冠状病毒S1蛋白,经超声裂解细菌后,取上清液进行亲和层析以纯化S1蛋白,并冻干浓缩;用纯化的S1蛋白免疫小鼠3次,间接酶联免疫吸附法(ELISA)检测产生的抗体IgG及其亚类的滴度,分析S1蛋白诱导的免疫应答。结果表达的S1蛋白主要存在于包涵体中,裂解上清液中存在少量S1蛋白;亲和层析纯化获得了能被SARS患者混合血清识别的S1蛋白,融合蛋白免疫小鼠3次后诱导产生了高滴度的IgG抗体和Th1/Th2型的免疫应答。结论纯化的S1蛋白具有良好的免疫学活性。

Objective To express and purify SARS Coronavirus spike protein fragment 1(S1) and analyze the immune response induced by S1. Methods E.coli JM109 containing recombinant plasmid expressing S1 protein was induced by IPTG to express S1 protein. Recombinant S1 was purified from supernatant of induced E.coli JM109 lysate containing recombinant S1; NIH mice were immunized with purified S1 on week 0, 2 and 6, antibody titer and their subtype induced by S1 was detected using indirect ELISA with S1,when the antibody titer of S1 was higher than or equal to 1∶6400, the mice were killed to collect the sera. Results Expressed S1 protein mainly existed in inclusion body and only small part in the supernatant. S1 was purified from E.coli lysate and the recombinant protein was reacted to mixed sera of ten SARS patients. Furthermore, mice immunized with S1 induced antibodies with a titer of 1∶25 600 and type Th1/Th2 immune response. Conclusion Purified S1 protein showed good immunological activity.