一种新的人精子特异性抗原表位肽的鉴定及意义

Identification and significance of a novel human sperm specific epitope peptide

目的从应用噬菌体展示随机肽库技术和生物信息学方法筛选出的候选人精子特异性抗原表位肽中鉴定新的人精子特异性抗原表位。方法多肽固相合成法合成候选人精子特异性抗原表位肽单体和三聚体。斑点免疫印迹法和酶联免疫吸附试验(ELISA)检测与抗精子抗体(ASA)阳性血清的免疫反应。ELISA检测免疫小鼠血清抗候选人精子特异性抗原表位肽抗体滴度。计算机辅助精子分析检测候选人精子特异性抗原表位肽免疫小鼠血清对人精子的作用。结果高效液相色谱法和质谱法测定结果显示,合成的候选人精子特异性抗原表位肽纯度和结构与设计相符。斑点免疫印迹法和ELISA检测结果显示,氨基酸序列为n-TRRIHRHMTIRR-c的候选人精子特异性抗原表位肽及其三聚体与35例ASA阳性血清中的17例呈较强的免疫反应。该候选人精子特异性抗原表位肽及其三聚体免疫小鼠血清处理人精子后,精子活力相关参数较对照组均明显降低(P<0.05),其中三聚体导致的精子活力相关参数下降尤为明显(P<0.01)。结论氨基酸序列为n-TRRIHRHMTIRR-c的十二肽具有人精子特异性抗原表位,是一种新的人精子特异性抗原表位肽。

Objective To screen and identify new human sperm specific antigen epitopes from candidate sperm-specific antigen epitope peptides using phage display random peptide library technology and bioinformatics methods.Methods Candidate sperm-specific epitope peptide monomers and trimers were synthesized by solid phase polypeptide synthesis.Dot immunoblotting and enzyme-linked immunosorbent assay(ELISA)were used to detect the immune reaction of the peptides with anti-sperm antibody(ASA)positive serum.ELISA was used to detect antibody titers against candidate sperm-specific epitope peptides in serum of immunized mice.The effects of the sera of the mice immunized with the candidate sperm-specific epitope peptide on human spermatozoa were examined by computer aided sperm analysis.Results High performance liquid chromatography and mass spectrometry showed that the purity and structure of the synthesized candidate sperm-specific epitope peptide were in accordance with the design.Blot and ELISA results indicated that the sperm-specific epitope peptide with the amino acid sequence n-TRRIHRHMTIRR-c and its trimers displayed strong immune response to 17 of 35 ASA positive sera.The serum from mice immunized with the peptide and its trimers significantly attenuated the sperm motility parameters compared with serum from the control group(P<0.05),with trimeric-induced decrease in motility parameters being particularly significant.(P<0.01).Conclusion The dodecapeptide with amino acid sequence n-TRRIHRHMTIRR-c is a novel human sperm-specific epitope peptide.