鹅细小病毒VP3蛋白的原核表达及纯化

Prokaryotic expression and purification of VP3 protein of goose parvovirus

目的原核表达并纯化鹅细小病毒(Goose parvovirus,GPV)VP3蛋白。方法将GPV VP3基因重组表达质粒pGEX-6P-VP3转化入大肠杆菌BL21(DE3)plysS内,IPTG诱导表达;用Sepharose 4B(Prepacked Glutathion)亲合层析柱纯化可溶性和不溶性包涵体蛋白,并进行SDS-PAGE、Western blot及Dot-ELISA鉴定。结果表达的GST/VP3融合蛋白相对分子质量为85 000,主要以不溶性包涵体形式存在;纯化的可溶性蛋白和包涵体蛋白含量分别为2.20和3.30 mg/ml;纯化的GST/VP3融合蛋白能被鹅抗GPV多克隆抗体特异性识别。结论原核表达并纯化了GPV VP3蛋白,为进一步研究其功能和免疫学特性及研制基因工程亚单位疫苗和新型抗原制剂奠定了基础。

Objective To express the VP3 protein of goose parvovirus （GPV） in prokaryotic cells and purify the expressed product. Methods Recombinant plasmid pGEX-VP3 was transformed into E. coli BL21 （DE3） plysS and induced with IPTG. The obtained soluble protein and inclusion body were purified by Sepharose 4B （Prepacked Glutathione） affinity chromatography, and i- dentified by SDS-PAGE, Western blot and Dot-ELISA. Results The expression fusion protein GST/VP3, with a relative molecular mass of 85 000, mainly existed in a form of insoluble inclusion body. The contents of purified soluble protein and inclusion body were 2. 20 and 3. 30 nag/ml respectively. Purified fusion protein GST/VP3 was specifically recognized by goose anti-GPV polyclonal antibody. Conclusion VP3 protein of GPV was expressed in prokaryotic cells and purified, which laid a foundation of further study on its function and immunological characters and development of recombinant subunit vaccine and novel antigen preparation.