title: Fluorescence and Absorbance Spectroscopy Methods to Study Membrane Perturbations by Antimicrobial Host Defense Peptides.
abstract: Antimicrobial peptides (AMPs) are currently intensely studied because of their potential as new bactericidal and bacteriostatic agents. The mechanism of action of numerous AMPs involves the permeabilization of bacterial membranes. Several methods have been developed to study peptide-membrane interactions; in particular optical spectroscopy methods are widely used. The intrinsic fluorescence properties of the Trp indole ring in Trp-containing AMPs can be exploited by measuring the fluorescence blue shift and acrylamide-induced fluorescence quenching. One important aspect of such studies is the use of distinct models of the bacterial membrane, in most cases large unilamellar vesicles (LUVs) with different, yet well-defined, phospholipid compositions. Deploying LUVs that are preloaded with fluorescent dyes, such as calcein, also allows for the study of vesicle permeabilization by AMPs. In addition, experiments using genetically engineered live Escherichia coli cells can be used to distinguish between the effects of AMPs on the outer and inner membranes of gram-negative bacteria. In combination, these methods can provide a detailed insight into the mode of action of AMPs.
authors.affiliation: Biochemistry Research Group, Department of Biological Sciences, University of Calgary, 2500 University Dr. NW, Calgary, AB, Canada, T2N 1N4.
authors.forename: Mauricio
authors.initials: M
authors.lastname: Arias
authors.affiliation: Biochemistry Research Group, Department of Biological Sciences, University of Calgary, 2500 University Dr. NW, Calgary, AB, Canada, T2N 1N4. vogel@ucalgary.ca.
authors.forename: Hans J
authors.initials: HJ
authors.lastname: Vogel
mesh_terms: D023181:Antimicrobial Cationic Peptides / Q000737:chemistry* / Q000378:metabolism; D002462:Cell Membrane / Q000737:chemistry* / Q000378:metabolism; D002463:Cell Membrane Permeability; D004926:Escherichia coli; D008563:Membrane Lipids / Q000737:chemistry / Q000378:metabolism; D010743:Phospholipids / Q000737:chemistry / Q000378:metabolism; D013050:Spectrometry, Fluorescence / Q000379:methods; D014364:Tryptophan; D053835:Unilamellar Liposomes / Q000737:chemistry / Q000378:metabolism; D056928:X-Ray Absorption Spectroscopy / Q000379:methods
chemical_list: D023181:Antimicrobial Cationic Peptides; D008563:Membrane Lipids; D010743:Phospholipids; D053835:Unilamellar Liposomes; D014364:Tryptophan
keywords: Absorbance spectroscopy; Antimicrobial peptides; Escherichia coli; Fluorescence spectroscopy; Large unilamellar vesicles; Membrane permeabilization; Tryptophan