国家自然科学基金(30130190),北京市自然科学基金(7012007)和北京大学肿瘤学重点学科资助项目.
This work was supported by grants from The National Natural Sciences Foundation of China (30130190), Natural Science Foundation of Beijing(7012007) and Oncology Key Program of Peking University.
既往工作表明,胃癌组织中有较高的猪鼻支原体感染率,猪鼻支原体的主要膜蛋白P37能够诱导外周血单核细胞释放肿瘤坏死因子(TNF).为了深入研究P37的作用机制,利用酵母双杂交系统筛选与P37蛋白相互作用的蛋白质分子.首先,将编码P37全长的cDNA克隆到pGBKT7载体中,构建“诱饵”表达载体pGBKT7-p37,以此筛选人胎盘组织cDNA表达文库.在2.6×106个克隆中,筛选到一株能与P37相互作用的阳性克隆,序列测定表明,该阳性克隆编码人视网膜色素上皮细胞蛋白(Norpeg蛋白).在此基础上经GST-Pull Down实验,进一步证实Norpeg蛋白确能与P37相互作用,为进一步研究P37对细胞的作用机制奠定了基础.
High ratio infection of Mycoplasma hyorhinis in gastric tumor tissues suggests a possible association between mycoplasma infection and cancer. P37, an outer-membrane bacterial protein from Mycoplasma hyorhinis, increases neoplastic invasivity and metastasis and induces TNF-α secretion from human peripheral blood mononuclear cells (PBMC). To investigate the functional mechanism of P37, yeast two-hybrid system was used to isolate proteins interacted with P37 from a human placenta cDNA library. Among the 2.6×106 transformant clones, one positive clone was obtained. Sequence analysis revealed that the clone is a cDNA fragment from Norpeg and encode a carboxy terminal protein. The interaction between P37 and Norpeg was further confirmed by ELISA and GST-Pull down assay. This result would be useful to study the function of P37 further.
孙玉宁,靳更林,张建芝,吴健,寿成超.猪鼻支原体P37蛋白相互作用蛋白的筛选与鉴定[J].生物化学与生物物理进展,2004,31(10):902-906
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