VEGFⅡ/GRP融合蛋白的构建、表达、纯化及其抗小鼠RM-1前列腺癌的作用研究

doi:10.13523/j.cb.20160802

中国生物工程杂志

2016, Vol. 36

Issue (8): 9-15 DOI: 10.13523/j.cb.20160802

研究报告

VEGFⅡ/GRP融合蛋白的构建、表达、纯化及其抗小鼠RM-1前列腺癌的作用研究

曹荣月, 俞敏霞, 张昕黎, 李曼曼, 苗梓韬, 金亮

中国药科大学生命科学与技术学院南京 210009

Construction,Expression,Purification of VEGFⅡ/GRP Fusion Protein and the Effects on RM-1 Prostate Tumor in Mice

CAO Rong-yue, YU Min-xia, ZHANG Xin-li, LI Man-man, MIAO Zi-tao, JIN Liang

School of Life Science and Technology, China Pharmaceutical University, Nanjing 210009, China

全文: PDF(892 KB) HTML

摘要：

设计并构建一种含VEGF和GRP抗原表位的表达载体pET28a-VEGFI-M2-GRP质粒，将其转化至大肠杆菌中，并对重组菌进行乳糖诱导表达，超声破碎，包涵体经洗涤、溶解、透析复性、离子交换层析等方法进行分离纯化后得到目的蛋白VEGFⅡ/GRP，Western blot 鉴定。建立前列腺癌RM-1细胞的C57BL/6J小鼠皮下移植瘤模型，以VEGFⅡ/GRP作为疫苗进行免疫。观察荷瘤小鼠的肿瘤生长情况计算抑瘤率，比较各组血管生成数以及ELISA检测抗VEGF抗体浓度，并研究该蛋白疫苗的抗肿瘤生长作用和抗血管生成作用。结果显示：ELISA结果表明小鼠血清中抗VEGF抗体比NS组高（P<0.05），重组蛋白VG组与NS组相比抗血管作用显著（P<0.05）。初步显示构建的重组蛋白有抑制肿瘤血管生长的作用。

关键词： VEGF; 融合蛋白; 抗血管生成; 前列腺癌; GRP

Abstract:

An expression vector pET-28a-VEGFⅠ-M₂-GRP plasmid containing the VEGF and GRP antigen epitopes was designed and constructed. After being expressed in recombined strains induced by lactose, the fusion protein was lysised by ultrasonic. The resulting inclusion body was washed,dissolved, and dialysised for renaturation and purified by ion-exchange chromatograph to get the target protein.VEGFⅡ/GRP was identified by Western blot. The C57BL/6J mice subcutaneous tumor model of prostate cancer cell line RM-1was established to receive treatments with VEGFⅡ/GRP vaccine.The tumor growth,the blood vessel densities and the concentration of anti-VEGF antibody were examined to assess the effect of anti-angiogenesis and anti-tumor. ELISA showed that the concentration of anti-VEGF antibody in the mice serum of VG group was significantly higher than that of NS group (P<0.05),the anti-angiogenesis effect of VG group was superior to that of NS group (P<0.05).Thus,the constructed fusion protein can inhibit angiogenesis.

Key words: Anti-prostate tumor Anti-angiogenesis VEGF Fusion protein GRP

收稿日期: 2016-02-01 出版日期: 2016-08-25

ZTFLH:

Q819

基金资助:

国家自然科学基金（81172973、81373232）、国家级大学生创新创业训练计划项目（J1030830，G14025）、江苏省高校优势学科建设工程（PAPD）、江苏省杰出人才引导项目（BK20140029）资助项目

通讯作者: 曹荣月, 金亮 E-mail: caorongyuenanjing@126.com;liangjin1975@cpu.edu.com

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引用本文:

曹荣月, 俞敏霞, 张昕黎, 李曼曼, 苗梓韬, 金亮. VEGFⅡ/GRP融合蛋白的构建、表达、纯化及其抗小鼠RM-1前列腺癌的作用研究[J]. 中国生物工程杂志, 2016, 36(8): 9-15.

CAO Rong-yue, YU Min-xia, ZHANG Xin-li, LI Man-man, MIAO Zi-tao, JIN Liang. Construction,Expression,Purification of VEGFⅡ/GRP Fusion Protein and the Effects on RM-1 Prostate Tumor in Mice. China Biotechnology, 2016, 36(8): 9-15.

链接本文:

https://manu60.magtech.com.cn/biotech/CN/10.13523/j.cb.20160802 或 https://manu60.magtech.com.cn/biotech/CN/Y2016/V36/I8/9

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