采用毛细管区带电泳研究人趋化素样因子1_C端活性多肽与肝素的相互作用.doc

采用毛细管区带电泳研究人趋化素样因子1 C端活性多肽人趋化素样因子1 C端活性多肽(C以不同比率混合温育 20min后进样，分离在6min内完成。结果 CKLF1-C19和CKLF1-C27与肝素均发生了相互作用，证实了最初的推断。通过Scatchard分析计算，得到CKLF1-C19和CKLF1-C27与肝素的结合常数分别为(3.38(0.49)×105M-1和(1.10(0.02)×105M-1。同时证明，CKLF1-C19肽链上的赖氨酸残基在相互作用的过程中发挥了重要作用，脯氨酸残基也会影响CKLF1-C19和肝素间的结合。 结论 本实验采用的毛细管区带电泳法研究CKLF1-C19和CKLF1-C27与肝素相互作用的方法快速、准确、可靠。为进一步探讨CKLF1-C19和CKLF1-C27在体内的活性及其生物学意义打下了基础。 关键词：毛细管区带电泳；CKLF1；肝素；肽；相互作用 Using capillary zone electrophoresis to study the interactions between the peptides from secreted human CKLF1 and heparin Yi Liu1, Shuyu Zhang2, Xiaomei Ling2, Yuzhen Li1, Yingmei Zhang3, Wenling Han3, Ying Wang3, 1. Department of Pharmacy, Peking University People’s Hospital, Beijing 100044, PR China 2. Department of Pharmaceutical Analysis, School of Pharmaceutical Sciences, Peking University, Beijing 100083, PR China 3. Center for Human Disease Genomics, Peking University, Beijing 100083, PR China ABSTRACT: OBJECTIVE Using a selective high-performance analytical method based on capillary zone electrophoresis (CZE) to investigate interactions between heparin and CKLF1-C27/ CKLF1-C19. Meanwhile, Using CZE to study the structural requirements of heparin and CKLF1-C19 recognition. METHODS Samples containing CKLF1-C27/CKLF1-C19/CKLF1-C19 km/CKLF1-C19pm and heparin at various ratios were incubated at room temperature and then separated by CZE with Tris-acetate buffer at pH 7.2. Each run was completed within 6 min. RESULTS Both qualitative and quantitative characterizations of the binding were determined. The binding constants of the interactions between CKLF1-C27/ CKLF1-C19 and heparin were calculated as (3.38(0.49)(105?M?1 and (1.10(0.02) ×105?M?1 by Scatchard analysis. We have found that the Pro or Lys to Ala substitution within the residues of CKLF1-C19 (CKLF1-C19pm or CKLF1-C19km) strongly decreased or abolished its interaction with heparin, suggesting that the residues of Pro affect the affinity of CKLF1-C19 for heparin, and the residues of Lys of CKLF1-C19 play the im