摘要
目的：获取原核表达的新型热休克蛋白分子HSP70L1与模型抗原卵清蛋白(OVA)片段的融合蛋白,并观察其生物学功能。方法：构建表达载体pQE30-OVA-HSP70L1,在相应的表达宿主菌中诱导表达后,从包涵体中依次以Ni^(2+)螯合层析柱和DEAE层析柱纯化目的蛋白；通过获得的融合蛋白对树突状细胞的成熟和细胞因子释放的促进作用初步检测其生物学活性。结果：pQE30-OVA-HSP70L1载体构建正确；纯化的OVA-HSP70L1融合蛋白相对分子质量约68 000,纯度大于95%；活性研究表明,OVA-HSP70L1融合蛋白可以有效地促进树突状细胞的成熟和Th1型细胞因子IL-12和TNF-α的产生。结论：纯化的HSPTOL1与抗原OVA融合蛋白具有一定的生物学功能,融合蛋白的获取为进一步开展HSP70L1佐剂效应机制研究奠定了基础。
Objective： To prepare, purify the recombinant proteins of HSP＇/0-1ike protein 1 （HSP70L1） with a large fragment of chicken ovalbumin （OVA） , and to investigate the bio-function of the fusion protein, providing a basis for further study of the effect and the mechanism of HSP70L1 as an adjuvant. Methods： The vector containing HSP70L1 cDNA and large fragment of OVA was constructed. The expression of OVA-HSP70L1 fusion protein was induced and the products were purified from inclusion bodies by His-Trap metal chelation chromatography and DEAE ion-exchange chromatography. The bio-activity of the fusion protein was examined by detecting its ability to activate dendritic ceils and to promote the secretion of cytokines. Results： The vector was successfully constructed and the molecular weight of the fused OVA- HSP70L1 protein （ with a purity of over 95% ） was 68 000. The fusion protein effectively promoted the maturation of dendritic cells and the production of cytokines such as interleukin-12 and tumor necrosis factor-α. Conclusion： HSP70L1 may be an effective adjuvant in the fusion protein and it may also promote antigen specific Thl type i mmol/Luno-responses.
出处
《中国肿瘤生物治疗杂志》
CAS
CSCD
2006年第2期129-134,共6页
Chinese Journal of Cancer Biotherapy
基金
全军医药科研"十五"规划重大项目(01Z058)
上海市科委重点科技攻关项目(044319208)
关键词
热休克蛋白
融合蛋白
树突状细胞
佐剂
heat shock protein
fusion protein
dendritic cell
adjuvant
