Protamine's action against Escherichia coli depends upon outer membrane structure and electrostatic interactions

Protamine is a cationic antimicrobial peptide being developed as a food preservative. The research presented in this thesis examines the effect of Escherichia coli outer membrane (OM) structure and charge on protamine sensitivity, and evaluates the roles of electrostatic interactions and inner core lipopolysaccharide (LPS) phosphates on protamine action. Protamine minimum inhibitory concentrations (MICs) ranged from 78--1250 mug/mL for 28 strains representing different OM structures. Smooth E. coli strains were up to 4x more resistant than rough strains. O-antigen was protective against protamine's initial OM-disrupting action, but had no apparent effect on cytoplasmic membrane permeabilization after long-term exposure. Smooth strains exhibited greater growth inhibition than rough strains over the first 8 hours of treatment at 2x MIC, due to increased clumping of rough cells. Loss of inner core phosphates reduced protamine susceptibility, altered protamine binding, and diminished the inhibitory effects of Na+ and Ca2+ ions on protamine action. These results confirm the importance of electrostatic interactions between protamine and the Gram negative OM.