Design of Novel Saposin-like Bacteriocins Using a Hybrid Approach

A multitude of approaches will be required to respond to the threat posed by the emergence and spread of antibiotic resistant pathogens. Bacteriocins have gained increasing attention as a possible alternative to antibiotics, as such peptide antimicrobials have mechanisms of action different from antibiotics and are therefore equally potent against antibiotic resistant bacteria as their susceptible counterparts. A group of bacteriocins known as saposin-like bacteriocins is believed to act directly on the bacterial membrane. Based on seven saposin-like leaderless bacteriocins, we have constructed a library of hybrid peptides containing all combinations of the N- and C-terminal halves of the native bacteriocins. All hybrid peptides were synthesized using in vitro protein expression and assayed for antimicrobial activity towards several pathogens. Of the 42 hybrid peptides, antimicrobial activity was confirmed for 11 novel hybrid peptides. Furthermore, several of the hybrid peptides exhibited altered antimicrobial spectra and apparent increase in potency compared to the peptides from which they were derived. The most promising hybrid, termed ISP26, was then obtained synthetically and shown to inhibit most of the Gram-positive species tested, including opportunistic pathogens and food spoilage bacteria. Additionally, ISP26 was shown to inhibit Acinetobacter, a species of Gram-negative bacteria frequently isolated from nosocomial infections. The activity of the hybrid library provides valuable insights into the design and screening of new active bacteriocins.