Methods: GL13K was incubated with the model Gram negative bacteria E.coli for 2-24 hours in 10 mM sodium phosphate pH 7.4, NaP+150 mM NaCl, PBS, 50% saliva in PBS or lysozyme (40-100mg/L) in PBS. Bactericidal activity was quantitated by enumerating the surviving colony forming units (CFU) or ATP content of live cells. Bacterial lysis was quantitated spectrophotometrically at 600nm. In some experiments the peptide was preincubated in water for 72 hours at room-temperature prior to testing activity. GL13K was compared to shorter C-terminal peptide KL7K to determine the functional domain.
Result: GL13K (10 or 100mg/L) killed 100% of E.coli in 10 mM NaP, NaP + 150mM NaCl, PBS or 50% saliva in PBS (similar to control Polymyxin B). GL13K retains antibacterial activity after incubation for 72 hours in water. Saliva caused lysis of GL13K and Polymyxin B-treated E.coli. This effect was partially replicated by lysozyme (40 or 100 mg/L). KL7K (C-terminal domain of GL13K) did not show an antibacterial effect.
Conclusion: The PSP peptide GL13K exhibits anti-inflammatory and antibacterial activity in the presence of physiological salt concentrations and saliva. Saliva enhances the lysis of E.coli in the presence of GL13K. The peptide retains activity in water, suggesting that it can be administered in drinking water. This suggests that GL13K is an attractive lead compound for development of oral antibiotics based on a human host-defense protein. Supported by NIH grant R01 DE017989.