噬菌体展示短肽模拟脂多糖结合蛋白抗炎功能位点的研究

Screening of peptide mimicking antiinflammatory functional site of lipopolysaccharide binding protein by phage peptide library

目的 应用噬菌体随机 12肽库筛选脂多糖结合蛋白 (lipopolysaccharidebindingprotein ,LBP)具有抗炎作用的肽序列。方法 以LPS为目标分子 ,对噬菌体十二肽库进行亲合筛选 ,4轮筛选后 ,检测随机挑取的噬菌体克隆的结合活性和抑制活性 ,对可与LBP竞争性结合LPS的噬菌体克隆行细胞因子生成抑制实验和LPS内化抑制实验 ,对获得的阳性克隆进行DNA序列测定 ,推导外源肽氨基酸序列 ,并与LBP序列比较 ,确定LBP的抗炎功能位点。结果 随机挑取的 10 0个噬菌体克隆中 16个可与LBP竞争性结合LPS ,其中 7个噬菌体克隆对LBP的增敏LPS刺激PBMC分泌TNF α功能无作用 ,对这 7个克隆进行LPS内化抑制实验 ,其中 3个克隆可以抑制LBP增强LPS内化作用。测序发现这 3个阳性克隆融合多肽的序列均为FHTRWNYWPYLH ,与LBP一级结构氨基酸序列同源性低。结论 该

Objective To screen mimic peptide of antiinflammatory functional site of lipopolysaccharide binding protein (LBP) from phage random 12 mer peptide library. Methods Using LPS as the target molecule, 4 rounds of bio panning to a phage random 12 mer peptide library were carried out. A total of 100 clones were selected and used in binding test, competitive inhibition test, and function test. Positive clones were sequenced and compared with LBP sequence. Results Sixteen phage clones out of the 100 clones could inhibit the binding of LBP to LPS, and 7 out of 16 clones had no effects on the function of LBP which enhanced the LPS dependent activation of PBMC. Three out of the 7 clones could inhibit the function of LBP to enhance LPS internalization, and had a consensus peptide sequence FHTRWNYWPYLH. Conclusion These results suggest that 12 mer peptide could mimic the antiinflammatiory functional site of LBP.