A Thermodynamic Study of the Effects of cholesterol on the Activity of Antimicrobial Peptide Protegrin-1

Abstract

Antibiotic resistance is a growing global concern. Whether by the unrestricted availability and overuse of antibiotics in the developing world or the tendency in the developed world to stop following antibiotic treatments before their prescribed end date, infectious bacterial strains have over time become resistant to these same antibiotics. A promising class of innate immune system molecules known as antimicrobial peptides (AMPs) may be a model for the next generation of antibiotic therapeutics. The most important characteristic of AMPs is their propensity to selectively attack and lyse bacterial membranes, while leaving mammalian (i.e., host) cell membranes intact. One of the critical differences between bacterial and mammalian cell membranes is the absence of cholesterol in the former and the sizable presence of cholesterol in the latter. Given that cholesterol stiffens a mammalian cell membrane above the gel-to-liquid phase transition temperature of its constituent phospholipids, we hypothesize that AMP selectivity is in part the result of the differing fluidities of mammalian and bacterial cell membranes. In our thermodynamic investigation of the relationship between membrane fluidity and the extent of AMP-induced membrane disruption, we have carried out isothermal titration calorimetry (ITC) measurements. Large unilamellar vesicles (LUVs) composed of varying ratios of 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) to cholesterol served as model membranes of varying fluidity, while the 18-residue long, cationic peptide protegrin-1 (PG-1) served as the AMP. In concert with previously-obtained atomic force microscopy (AFM) data, our ITC results confirm that cholesterol indeed protects phospholipid membranes from AMP-induced disruption.