Abstract
Some chemokines have been shown act as antimicrobial proteins in vitro, suggesting that they may contribute directly to innate immune protection. Based on the study of defensins and other antimicrobial peptides (AMPs), it has been proposed that the characteristics of cationicity, hydrophobicity and amphipathicity of the AMPs determine the killing activity of these peptides. However, amino acid characteristics necessary for the antimicrobial activity of chemokines have not been elucidated. Using mouse CCL28 we have determined the primary sequence features required for antimicrobial activity of this chemokine. Our data demonstrate, through the use of successive truncations, that the holoprotein (108 amino acids) is necessary for full antimicrobial activity of CCL28. Consecutive truncations resulted in an almost complete loss of activity following the removal of the 24 C terminal amino acids. Charge reversal as well as positive charge to neutral charge mutations, support the hypothesis that positively charged amino acids are essential for the antimicrobial activity of CCL28. Finally, the antimicrobial activity of chimeric proteins in which the N-terminus of CCL5 or CCL27 was fused with the C terminus of CCL28 suggest that although the C terminal region of CCL28 is essential for antimicrobial activity, the interaction of the N and C terminal regions of CCL28 is necessary for optimal antimicrobial activity. These experiments are the first to investigate the role of primary structure in the killing activity of an antimicrobial chemokine.
