摘要
用RTPCR方法从人单核THP1细胞系克隆人41BBL胞外区基因,将其重组到pAYZ表达载体中,构建成人41BBL胞外区基因表达载体。将该载体转化大肠杆菌16C9,获得稳定表达,表达产物主要以可溶性状态存在;SDSPAGE和Westernblot分析显示,其分子量约为22kD,与预期结果一致。这是首次在大肠杆菌中获得41BBL胞外区可溶性表达。生物学活性检测显示41BBL对于维持T淋巴细胞系因子释放非常有益,同时PI单染表明它能抑制Jurkat细胞的凋亡。这将在抗肿瘤免疫治疗中具有潜在应用前景。
RT-PCR was used to clone DNA fragment of the extracellular domain of 4-1BBL from human THP-1 cells （human monocyte）, and the expression vector pAYZ4-1BBL was constructed by cloning the extracellular domain of 4-1BBL into the expression vector pAYZ. The extracellular domain of 4-1BBL was expressed in E. coli 16C9 and purified by affinity chromatography, SDS-PAGE and Western blot analysis showed that the relativae molecular weight of soluble 4-1BBL is 22kD which was consistent with the theoretically predicted value. So far as we know, it is the first time that the soluble expression of 4-1BBL in E. coli.was achieved 4-1BBL induced a significant release of IL-2 in stimulated Jurkat cells after 48h incubation, especially in the presence of tumor cell. At the same time the apoptosis level of Jurkat cell reduce more than 50%. In conclusion,4-1BBL may be useful in cancer immunotherapy.
出处
《生物工程学报》
CAS
CSCD
北大核心
2005年第5期703-707,共5页
Chinese Journal of Biotechnology
基金
国家高技术研究发展计划项目(863)资助(No.2003AA215080)
天津市重大科技攻关经费资助(No.003119511)。~~
关键词
4-1BBL
协同刺激分子
原核表达
4-1BB ligand, costimulatory molecules, prokaryotic expression
