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第三军医大学西南医院病理学研究所
纸质出版日期:2008,
网络出版日期:2008-12-30,
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杨海捷, 段光杰, 刘友生, 等. 运用噬菌体展示随机肽库筛选与内毒素结合的高亲和性多肽[J]. 陆军军医大学学报, 2008,30(24):2262-2266.
YANG HAI-JIE, DUAN GUANG-JIE, LIU YOU-SHENG, et al. High affinity peptides binding to LPS screened from phage display random peptide library. [J]. Journal of army medical university, 2008, 30(24): 2262-2266.
目的 从噬菌体展示随机肽库中筛选与内毒素结合的多肽序列,并进行鉴定。 方法 以内毒素脂多糖(lipopolysaccharide, LPS)为靶分子对噬菌体展示随机十二肽库进行4轮亲和筛选,获得与LPS结合的噬菌体克隆,应用结合实验和克隆斑抑制实验进一步确证。挑选结合力强的克隆进行DNA测序,推导出呈现的多肽序列,应用生物信息学软件进行多肽序列分析和同源性分析。 结果 经4轮亲和筛选从噬菌体展示随机十二肽库中筛选获得了86个克隆,挑选12个结合力强的克隆进行DNA序列测序及生物信息学分析,结合本项目组的噬菌体展示随机七肽库筛选结果推导出呈现的多肽序列为HWQWPHWSPPP(命名为P11肽)。检索相关数据库发现此肽序列未被申请专利,体内约908种蛋白与其结构相匹配,其中包含有与LPS相互作用的位点。 结论 通过对噬菌体展示随机肽库的淘选,获得与LPS结合的高亲和性多肽,为进一步以这些多肽为先导物进行定向进化研究提供了实验依据和结构基础。
Objective To screen and identify peptides with high affinity to lipopolysaccharide (LPS) from phage display random peptide library. Methods Using LPS as a ligand
phage clones that bond to LPS were screened from phage display random 12-mer peptide library through 4 rounds of biopanning. After binding activity was measured by ELISA and plaque reduction testing
the phage clones presenting high affinity to LPS was selected and amplified. Their DNA were extracted for sequencing
and the coding peptide sequences were deduced from the results and analyzed by bioinformatics including homology analysis through blasting. Results After 4 rounds of effective screen
86 eluted clones were obtained and amino acid sequences of 12 positive clones with high affinity to LPS were selected and identified. The core peptide sequence of phage clones was deduced as HWQWPHWSPPP (named P11) through comparing with the results from phage display random 7-mer peptide library
which was considered to contain the motif of the common amino acid sequence for LPS affinity. There was no patent declared based on P11 sequences retrieving from patent database. There were 908 proteins matching partly to the peptide found by homology retrieval. Conclusion Peptides with high affinity to LPS are obtained from phage display random peptide library
which provide us structural foundations on further study of direct-evolution using these peptides as primers.
内毒素噬菌体展示随机肽库多肽生物信息学
lipopolysaccharidephage display random peptide libraryPeptidebioinformatics
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