Abstract
The Chemokine‐like factor 1 (CKLF1) is a novel human cytokine and exhibits chemotactic activities on leukocytes. Two peptides named CKLF1‐C27 and CKLF1‐C19, were obtained from secreted CKLF1. In this study, a selective high‐performance analytical method based on capillary zone electrophoresis (CZE) to investigate interactions between heparin and CKLF1‐C27/CKLF1‐C19 was developed. Samples containing CKLF1‐C27/CKLF1‐C19 and heparin at various ratios were incubated at room temperature and then separated by CZE with Tris‐acetate buffer at pH 7.2. 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) × 10^5^ M^−1^ and (1.10 ± 0.02) × 10^5^ M^−1^ by Scatchard analysis. To study structural requirements, CKLF1‐C19pm and CKLF1‐C19km have been synthesized, and their interactions with heparin have been studied by CZE. We 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 important role for the interaction of CKLF1‐C19 and heparin, respectively. The methodology presented should be generally applicable to study peptides and heparin interactions quantitatively and qualitatively. Copyright © 2008 European Peptide Society and John Wiley & Sons, Ltd.