2H Solid-State NMR Studies of the Antimicrobial Peptide MSI-78 Interacting with the Membranes of Whole Escherichia Coli

Abstract

Antimicrobial peptides (AMPs) are ubiquitous molecules that can display antimicrobial activity against bacteria, viruses, protozoa and various other pathogens. A key aspect of AMP activity is their interactions with biological membranes. Solid-state NMR spectroscopy has thus been an important tool for their study. Previous experiments, on model membrane systems, have elucidated important aspects of AMP mechanism. However, the extent to which actual in vivo AMP activity can be understood from model studies is necessarily limited. Peptide-membrane interactions under physiological conditions are presumably influenced by additional factors such as: interactions with lipopolysaccharides, the presence of membrane proteins, membrane compositional heterogeneity, lipid domains, etc. In order to bridge the gap between the NMR studies of AMPs using model membranes and the AMP-membrane interactions occurring in intact cells, we have designed a procedure to incorporate high levels of 2H-NMR labels, specifically into the cell membrane, by creating a novel strain of E. coli: LA8. Using this strain we are able to reproducibly quantify the effects of the AMP MSI-78 on lipid chain order in bacterial membranes. Treatment with MSI-78 led to an increase in the disorder of the bacterial membrane. This was observed by the decrease in the average order parameter and by the increase in intensity at the lower frequencies. The peptide:lipid ratios needed to observe MSI-78's effects on acyl chain order in the intact cells falls between the ratios required to observe effects in NMR studies of model lipid systems and the ratios required to observe inhibition of cell growth in biological assays.