摘要
制备16型人乳头瘤病毒mE6Δ/mE7蛋白与I型人单纯疱疹病毒VP22Δ蛋白的治疗型分子内佐剂融合蛋白疫苗,并检测其免疫原性和抗肿瘤相关生物活性。通过克隆HSV-1 VP22Δ及HPV-16 mE6Δ/mE7基因,构建pET28a-VP22Δ-mE6Δ/mE7原核表达载体。重组质粒在Rosetta(DE3)宿主菌中进行诱导表达,表达蛋白经分离、复性后,通过镍离子亲和层析进行纯化,纯化蛋白经SDS-PAGE、Western blot鉴定,并免疫BalB/C及C57BL/6小鼠,检测其免疫原性和抗肿瘤活性。结果显示,VP22Δ-mE6Δ/mE7蛋白以包涵体形式表达,分子量约为34kDa,表达量约占菌体总蛋白的45%。该蛋白免疫小鼠后血清特异性IgG、特异性淋巴细胞增殖效果及对TC-1致瘤小鼠的肿瘤治疗效果均高于无佐剂单一重组蛋白疫苗。以上结果说明,所获得的重组融合蛋白具有较好的免疫原性和抗肿瘤活性,为治疗型HPV分子内佐剂疫苗的进一步研究奠定了基础。
In order to investigate the biological effects of the VP22Δ-mE6Δ/mE7 built-in adjuvant fusion protein vaccine on the tumor associated with HPV-16 infection. HSV-1 VP22A and HPV-16 mE6△/mE7 genes were cloned, and the pET28a-VP22△-mE6△/mE7 recombinat prokaryotic expression vector was constructed. Vector was transformed into Rosetta （DE3）E. coli string and expressed under the induction of IPTG. The renaturalized protein was then purified via Ni2 ＋ affinity adsorbent column and identified by SDS-PAGE and Western blot. Purified protein was immunized BalB/C and C57BL/6 mice to evaluate the immunogenieity and anti -tumor activity. The expressed recombinant protein formed as inclusion body with a prediction MW about 34kDa and contained approximately 45% of total somatic protein. The VP22△-mE6△/mE7 immunization induced higher titer of specific IgG against HPV, higher level of lymphocyte proliferation and better effect on suppressing HPV16 positive TC-1 tumor growth than the mice immunized with mE6△/mE7 alone. The results showed that the recombined built-in adjuvant vaccine could induce specific cellular immune response in vitro and inhibit the TC- 1 tumor proliferation in vivo, that would be a foundation for further studies and developments of inner adjuvant vaccines.
出处
《中国生物工程杂志》
CAS
CSCD
北大核心
2009年第4期6-11,共6页
China Biotechnology
基金
云南省自然科学基金资助项目(2007C144M)
