Effect of zinc and phosphate on an antibacterial peptide isolated from lung lavage

Incubation of Escherichia coli (10(4) organisms per ml) in cell-free rabbit lung lavage for 30 min at 37 degrees C resulted in a 70% reduction in colony counts on deoxycholate agar. A low-molecular-weight peptide (about 3,400 daltons), with zinc as a cofactor, was responsible for this activity. The peptide was isolated by Sephadex G-15 separation of lyophilized rabbit lung lavage which had been centrifuged to remove macrophages and suspended phospholipids and passed through a 10,000-dalton (pore size) membrane filter. Peptide activity against E. coli was inhibited by phosphate buffer but not by borate, Tris, or barbital buffer. Bacteria incubated in phosphate buffer and then washed in saline were resistant to peptide activity. Antibacterial activity was also inhibited when peptide-exposed bacteria were incubated in phosphate buffer before deoxycholate treatment. 32P-radiolabeled E. coli cells lost about 20% of their radiolabel after 15 min of incubation with peptide.