转化生长因子β1的关键序列筛选及鉴定

The Biopanning and Appraisal of Key Sequence of Transforming Growth Factor-beta 1

目的应用噬菌体随机12肽库,筛选转化生长因子β1(Transforming growth factor-beta1,TGF-β1)的关键序列,进行TGF-β1活性短肽的合成,并检测其与成纤维细胞的亲和力。方法以TGF-β1单克隆抗体为靶,筛选噬菌体随机12肽库,获得TGF-β1的特异性序列。进行序列比对分析,选择TGF-β1的关键序列。进行TGF-β1活性短肽的合成、纯化、修饰及免疫荧光标记。应用免疫荧光法检测TGF-β1活性短肽的成纤维细胞亲和力。结果筛选噬菌体随机12肽库,获得10个与TGF-β1相似的特异性序列。分析获得7个TGF-β1的关键序列。合成获得7个TGF-β1活性短肽,并纯化至98%,短肽的C端进行氨基化封闭,N端进行罗丹明激光染料标记。免疫荧光检测结果显示,1个TGF-β1活性短肽能够与成纤维细胞相结合。结论从噬菌体随机12肽库中筛选到TGF-β1的关键序列,1个TGF-β1活性短肽能够与成纤维细胞相结合,有望应用于促进创面愈合的相关研究。

Objective To isolate key sequences from a phage display 12-mer peptide library, synthesize transforming growth factor-beta1（TGF-β1） bioactive peptides and evaluate the fibroblasts affinity of TGF-β1 bioactive peptides.Methods Using monoclonal anti-human TGF-β1 antibody as the target, a phage display 12-mer peptide library was screened and target sequences were isolated. The key sequences of TGF-β1 were chosen. The key sequences were then modified to generate TGF-β1 bioactive peptides. Immunofluorescence assay was performed to evaluate the fibroblasts affinity of TGF-β1 bioactive peptides. Results Ten peptides similar to TGF-β1 were isolated from a phage display 12-mer peptide library. Seven key sequences of TGF-β1 were chosen and then modified to generate TGF-β1 bioactive peptides. TGF-β1bioactive peptides were then purified to reach a purity of 98%. The C terminal of TGF-β1 bioactive peptides was closed, and the N terminal was labeled with rhodamine dye laser. Immunofluorescence assay showed one TGF- β1 bioactive peptides was able to combine with the fibroblasts. Conclusion The key sequences of TGF-β1 can be screened from a phage display 12-mer peptide library. One TGF-β1 bioactive peptides can combine with the fibroblasts. The results are expected to help promoting wound healing.