抗HIV-1单链抗体重组白喉毒素的构建与表达研究

发布时间：2018-01-15 15:07

本文关键词：抗HIV-1单链抗体重组白喉毒素的构建与表达研究　出处：《长春理工大学》2007年硕士论文　论文类型：学位论文

【摘要】：重组免疫毒素被称为“生物导弹”，是目前肿瘤、自身免疫性疾病以及移植排斥等免疫治疗领域的热点研究方向之一。对于AIDS而言，由于免疫毒素能够特异性的杀伤HIV潜伏感染细胞，彻底摧毁病毒储存池，而对正常细胞毒性很低，从而成为当前HAART辅助治疗的亮点。为了获得针对HIV-1的无毒副作用且能根除体内病毒的药物，进行了重组免疫毒素的表达，期望其针对HIV病毒和感染细胞的特异杀伤作用可以最终实现抗病毒这一治疗目的。将获得的单链抗体(ScFv)基因分别代替白喉毒素DT的受体结合区以及白喉毒素DT的受体结合区和跨膜区，制备出重组毒素基因DT-ScFv，利用原核表达载体pET28a，构建了用于表达重组毒素的原核表达质粒pET-DT-hS120、pET-DTA-hS120，上述质粒转化受体菌BL21(DE3)后，经IPTG诱导，成功地表达了目的蛋白，表达量分别为21％，23％，，并利用Ni-NTA金属螯合层析法对表达产物进行了纯化及复性处理。
[Abstract]:Recombinant immunotoxin called "biological missile", is currently a hot research direction of tumor, autoimmune diseases and transplant rejection and other immune therapeutic areas. For AIDS, because the immune cell specific toxins can kill HIV virus latent infection, completely destroyed the storage tank and low toxicity to normal cells. In order to become the HAART adjuvant therapy highlights.
In order to get rid of the toxic side effects of HIV-1 and eradicate the virus in vivo, the expression of recombinant immunotoxin is carried out. It is hoped that its specific killing effect against HIV virus and infected cells can finally achieve the purpose of antiviral treatment.
The single chain antibody (ScFv) gene instead of diphtheria toxin DT receptor binding and receptor DT of diphtheria toxin binding region and transmembrane region, prepared recombinant toxin DT-ScFv gene, prokaryotic expression vector pET28a for expression of recombinant toxin, the prokaryotic expression vector pET-DT-hS120 pET-DTA-hS120 was constructed. The plasmid was transformed into recipient BL21 (DE3), induced by IPTG, the successful expression of the target protein, the expression level was 21%, 23% respectively, and the expression products were purified and renatured by Ni-NTA metal chelate affinity chromatography.

【学位授予单位】：长春理工大学
【学位级别】：硕士
【学位授予年份】：2007
【分类号】：R392

【参考文献】