Antimicrobial peptides are important effectors of the innate immune system. These peptides belong to a multifunctional group of molecules that apart from their antibacterial activities also interact with mammalian cells and glycosaminoglycans and control chemotaxis, apoptosis, and angiogenesis. Here we demonstrate a novel antimicrobial activity of the heparin-binding and cell-binding domain 5 of high molecular weight kininogen. Antimicrobial epitopes of domain 5 were characterized by analysis

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... overlapping peptides. A peptide, HKH20 (His 479 -His 498 ), efficiently killed the Gram-negative bacteria Escherichia coli and Pseudomonas aeruginosa and the Gram-positive Enterococcus faecalis. Fluorescence microscopy and electron microscopy demonstrated that HKH20 binds to and induces breaks in bacterial membranes. Furthermore, no discernible hemolysis or membrane-permeabilizing effects on eukaryotic cells were noted. Proteolytic degradation of high molecular weight kininogen by neutrophil-derived proteases as well as the metalloproteinase elastase from P. aeruginosa yielded fragments comprising HKH20 epitopes, indicating that kininogen-derived antibacterial peptides are released during proteolysis. . 2 The abbreviations used are: AMP, antimicrobial peptide; HMWK, high molecular weight kininogen; D5, domain 5; rD5, recombinant domain 5; RDA, radial diffusion assays; PBS, phosphate-buffered saline; LDH, lactate dehydrogenase; MEC, minimal effective concentration; cfu, colony-forming units; DMEM, Dulbecco's modified Eagle's medium; MALDI-TOF, matrix-assisted laser desorption ionization time-of-flight; MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; Tricine, N-[2-hydroxy-1, 1-bis(hydroxymethyl)ethyl]glycine.