Abstract: Two lactoferrampin (LFampin) peptides derived from bovine lactoferrin were compared with respect to their bactericidal activities. LFampin 265-284 killed a set of Gram-positive bacteria that were resistant to LFampin 268-284. The presence of [sup.265]Asp-Leu-[sup.267]Ile did not simply lead to an overall increased potency, since higher concentrations of LFampin 265-284 than LFampin 268-284 were needed to kill the Gram-negative bacteria that were tested. The Asp-Leu-Ile sequence enhances the propensity of LFampin to adopt an [alpha]-helix, as shown by circular dichroism spectroscopy. These results suggest that the helical conformation of the peptide is an important determinant of the susceptibility of Gram-positive bacteria.
Key words: [alpha]-helix, antimicrobial peptides, Gram-negative bacteria, Gram-positive bacteria, lactoferrampin, lactoferrin.
Resume : Deux peptides derives de la lactoferrine bovine appeles lactoferrampines (LFampin) ont ete compares quant a leurs activites bactericides respectives. La LFampin 265-284 a tue une variete de bacteries Gram-positives qui etaient resistantes a la LFampin 268-284. La presence des residus [Asp.sup.265]-Leu-[Ile.sup.267] ne resultait pas seulement en l'augmentation generale du potentiel bactericide du peptide, car la LFampin 268-284 tuait de facon plus efficace les bacteries Gram-negatives que la LFampin 265-284. La sequence Asp-Leu-Ile augmente la propension de la LFampin a adopter une structure en helice-[alpha], tel que demontre en spectroscopie par dichroisme circulaire. Ces resultats suggerent que la conformation helicoidale du peptide soit un determinant important dans la susceptibilite des bacteries Gram-positives.
Mots cles : helice-[alpha], peptides anti-microbiens, bacterie Gram-positive, bacterie Gram-negative, lactoferrampine, lactoferrine.
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Introduction
Antimicrobial peptides provide a rapidly expanding source of new antimicrobial compounds (Van 't Hof et al. 2001). They were first discovered in insects and rabbit neutrophils (Selsted et al. 1983; Steiner 1982). Subsequent findings of antimicrobial peptides in human, frogs, pigs, and cattle (Pollock et al. 1984; Lee et al. 1989; Tomita et al. 1991; Zasloff 1987) made it clear that antimicrobial peptides are widespread in nature. Some antimicrobial peptides originate from precursor proteins through proteolytic digestion (Bals and Wilson 2003; Tomita et al. 1991). Lactoferricin B is released from bovine lactoferrin by pepsin digestion and has antibacterial (Bellamy et al. 1992; Yamauchi et al. 1993), antifungal (Wakabayashi et al. 1996), and antiviral activity (Valenti et al. 2000). Related synthetic peptides show broad-spectrum bactericidal activities against a range of Gram-positive and Gram-negative bacteria, such as several streptococci, Escherichia coli, Bacillus subtilis, Staphylococcus aureus, Porphyromonas gingivalis, and Prevotella intermedia (Tomita et al. 1994; Kang et al. 1996; Groenink et al. 1999; Vorland et al. 1999; Gifford et al. 2005).
We have identified another antimicrobial peptide between residues 268 and 284 in the N1-domain of bovine lactoferrin, designated LFampin (Van der Kraan et al. 2004). This peptide displayed antimicrobial activity against the yeast Candida albicans and a number of bacteria, including E. coli, B. subtilis, and Pseudomonas aeruginosa. Using a series of peptides spanning the LFampin domain, we demonstrated that LFampin 265-284 possessed the highest candidacidal activity (Van der Kraan et al. 2005a). Unexpectedly, since net positive charge is one of the general features of antimicrobial peptides, the presence of the 3 extra amino acids [sup.265]Asp-Leu-[sup.267]Ile, including a negatively charged one, substantially increased the candidacidal activity. A peptide comprising the antimicrobial sequence could be functionally released from bovine lactoferrin by 1-enzyme digestion. The resulting isolated peptide, as well as the total digest, was …
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