采用CDR3亲和力转移方法快速获得抗FSHR抗体

发布时间：2018-01-23 03:22

本文关键词： 亲和力转移 FSHR FSH- c Ab BCII 纳米抗体　出处：《新疆农业科学》2017年07期 　论文类型：期刊论文

【摘要】：【目的】评价骆驼来源的纳米抗体c Ab BCII10作为非抗体亲和力转移骨架的潜能,采用亲和力转移的方法,将FSHR结合肽段移植到c Ab BCII10的抗原结合区以快速获得抗FSHR抗体。【方法】将FSHR结合肽FSH33-53编码序列,分别移植入纳米抗体c Ab BCII10的CDR1和CDR3区域中,命名为VHH-h FSH1和VHH-h FSH3。采用DNA合成方法获得VHH-h FSH1,VHH-h FSH3和c Ab BCII10的DNA编码序列,把这些DNA序列克隆到p ET22b载体上,将其转化到BL21(DE3)感受态细胞中。经过诱导和表达再用Ni离子亲和纯化获得纯的单域抗体。采取ELISA方法鉴定纯化的c Ab BCII10,VHH-h FSH1、VHH-h FSH3与FSHR的结合能力及特异性。【结果】通过框架移植获得的VHH-h FSH1,VHH-h FSH3和c Ab BCII10蛋白均在细菌胞间质可溶性表达。将FSHR结合肽FSH33-53移植到c Ab BCII10的CDR3获得的VHH-h FSH3具有特异结合FSHR活性。【结论】c Ab BCII10可以作为移植的框架,FSH和FSHR结合肽段移植到c Ab BCII10的CDR1和CDR3区可以获得亲和性较高的抗FSHR抗体。
[Abstract]:[objective] to evaluate the potential of camel-derived nano-antibody BCII10 as the skeleton of non-antibody affinity transfer, and to use affinity transfer method. The FSHR binding peptide fragment was transplanted into the antigenic binding region of BCII10 in order to obtain anti-#en2# antibody quickly. [methods] the coding sequence of FSHR binding peptide FSH33-53 was obtained. It was transplanted into the CDR1 and CDR3 regions of BCII10. Named VHH-h FSH1 and VHH-h FSH3. VHH-h FSH1 was obtained by DNA synthesis method. The DNA coding sequences of VHH-h FSH3 and c-Ab BCII10 were cloned into p ET22b vector. Convert it to BL21 DE3). The purified single domain antibody was obtained by elicitation and expression and then purified by Ni ion affinity. The purified BCII10 was identified by ELISA method. The binding ability and specificity of VHH-h FSH1VHH-h FSH3 to FSHR. [results] VHH-h FSH1 obtained by frame transplantation. VHH-h FSH3 and c Ab BCII10 protein were expressed in the interstitial cells of bacteria. The FSHR binding peptide FSH33-53 was transplanted into c Ab. VHH-h FSH3 obtained from CDR3 of BCII10 has specific binding FSHR activity. [conclusion] c Ab BCII10 can be used as a framework for transplantation. FSH and FSHR binding peptides were transplanted into the CDR1 and CDR3 regions of c Ab BCII10 to obtain anti FSHR antibodies with high affinity.
【作者单位】：新疆大学生命科学与技术学院/新疆生物资源基因工程省部共建重点实验室;
【基金】：国家自然科学基金项目“骆驼单域抗体框架区作为小分子抗体通用移植骨架的研究”(31370933);国家自然科学基金项目“基于高通量测序对骆驼抗体库的多样性分析和数据挖掘及新纳米抗体的发现”(31570935)~~
【分类号】：R392
【正文快照】： 0引言【研究意义】卵泡刺激素受体(follicle-stimu-lating hormone receptor,FSHR)是一种跨膜糖蛋白,属于G蛋白偶联受体超家族中糖蛋白亚家族成员。近期发现FSHR普遍表达于各种进程的肿瘤新生血管内皮的细胞表面。表明了FSHR是一种合适的肿瘤标志物和抗体作用的良好靶点。【前

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