卵泡刺激素功能肽与受体胞外区不同片段结合作用的分析

发布时间：2018-05-14 20:41

本文选题：FSH-肽 + LRR　；参考：《新疆农业科学》2017年07期

【摘要】：【目的】卵泡刺激素(FSH)33-53肽可以与FSH受体(FSHR)结合,并激活下游信号,被作为FSH功能肽。然而其在受体上的具体结合位置还不清楚。阐明FSH33-53肽在FSHR上的结合区域,为基于FSH的疫苗设计提供依据。【方法】采用重组的FSHR胞外区(ECD)和富含亮氨酸区(LRR)及合成的FSHR9-30区分析这些FSHR的功能区域与FSH33-53的结合作用。PCR扩增得到h FSHR ECD和h FSHR LRR目的基因,构建重组质粒p ET22b-FSHR-ECD和p ET22b-FSHR-LRR,表达纯化获得蛋白FSHR-ECD和FSHR-LRR。采用多肽合成法获得FSHR9-30-KLH。通过ELISA方法检测受体片段与FSH33-53肽的结合和亲和力。【结果】获得FSHR-ECD和FSHR-LRR蛋白,其相对分子质量(Mr)分别为43和32k Da。在受体为0.5μg湓m L,配体为2.5μg湓m L时,三种蛋白均与FSH33-53肽结合。ELISA检测受体与配体的亲和力分别为0.21×10~(-6)、0.45×10~(-6)和0.056×10~(-6)mol/L。【结论】FSHR 9-30片段与FSH33-53肽的结合较其它两个片段结合力强。
[Abstract]:[objective] FSH 33-53 peptide can bind to FSH receptor FSHR and activate downstream signal, and be used as FSH functional peptide. However, the exact site of its binding to the receptor remains unclear. To elucidate the binding region of FSH33-53 peptides on FSHR, [methods] Recombinant FSHR extracellular region (ECD) and leucine-rich region (LRR) and synthetic FSHR9-30 region were used to analyze the binding of these FSHR functional regions to FSH33-53. [methods] h FSHR ECD and h FSHR LRR target genes were obtained by PCR amplification. The recombinant plasmids p ET22b-FSHR-ECD and pET22b-FSHR-LRR were constructed, and the protein FSHR-ECD and FSHR-LRR were purified. FSHR9-30-KLH was synthesized by polypeptide synthesis. The binding and affinity of the receptor fragments to FSH33-53 peptides were detected by ELISA method. [results] FSHR-ECD and FSHR-LRR proteins were obtained with relative molecular weight of 43 and 32 k Da. respectively. When the receptor was 0.5 渭 g / mL and the ligand was 2.5 渭 g / mL, the binding ability of the three proteins to FSH33-53 peptide was 0.21 脳 10 ~ (-6) and 0.056 脳 10 ~ (10) ~ (6) mol 路L ~ (-1), respectively. [conclusion] the binding ability of FSHR 9-30 fragment to FSH33-53 peptide was stronger than that of the other two fragments.
【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重点实验室;
【基金】：国家自然科学基金项目“抗卵泡刺激素受体纳米抗体的制备及其在肿瘤分子显像和抗血管治疗中的作用”(81260333)~~
【分类号】：Q78;R91

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