Peptides of the C-terminal region of human
thrombin are released upon proteolysis and identified in human
wounds. In this study, we wanted to investigate minimal determinants, as well as structural features, governing the antimicrobial and immunomodulating activity of this
peptide region. Sequential
amino acid deletions of the
peptide GKYGFYTHVFRLKKWIQKVIDQFGE (GKY25), as well as substitutions at strategic and structurally relevant positions, were followed by analyses of antimicrobial activity against the Gram-negative bacteria Escherichia coli and Pseudomonas aeruginosa, the Gram-positive bacterium Staphylococcus aureus, and the fungus Candida albicans. Furthermore,
peptide effects on
lipopolysaccharide (LPS)-,
lipoteichoic acid-, or
zymosan-induced macrophage activation were studied. The
thrombin-derived
peptides displayed length- and sequence-dependent antimicrobial as well as immunomodulating effects. A
peptide length of at least 20
amino acids was required for effective anti-inflammatory effects in macrophage models, as well as optimal antimicrobial activity as judged by MIC assays. However, shorter (>12
amino acids) variants also displayed significant antimicrobial effects. A central K14 residue was important for optimal antimicrobial activity. Finally, one
peptide variant, GKYGFYTHVFRLKKWIQKVI (GKY20) exhibiting improved selectivity, i.e., low toxicity and a preserved antimicrobial as well as anti-inflammatory effect, showed efficiency in mouse models of LPS
shock and P. aeruginosa
sepsis. The work defines structure-activity relationships of C-terminal
host defense peptides of
thrombin and delineates a strategy for selecting
peptide epitopes of therapeutic interest.