Abstract
The antibiotic activity of antimicrobial peptides is generally derived via some type of disruption of the cell membrane(s). The most common models used to mimic the properties of bacterial membranes consist of mixtures of various zwitterionic and anionic phospholipids. This approach works reasonably well for Gram-positive bacteria. However, since the membranes of Gram-negative bacteria contain lipopolysaccharides, as well as zwitterionic and anionic phospholipids, a more complex model is required to simulate the outer membrane of Gram-negative bacteria. Herein we present a protocol for the preparation of models of the outer membranes of the Gram-negative bacteria Klebsiella pneumoniae and Pseudomonas aeruginosa. This protocol can be used to prepare models of other Gram-negative bacteria provided the strain-specific lipopolysaccharides are available.
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Bechinger B (2011) Insights into the mechanism of action of host defense peptides from biophysical and structural investigations. J Pept Sci 17:306–314
Brogden KA (2005) Antimicrobial peptides: pore formers or metabolic inhibitors in bacteria? Nat Rev Microbiol 3:238–250
Domadia PN, Bhunia A, Ramamoorthy A, Bhattacharjya S (2010) Structure, interactions, and antimicrobial activities of MSI-594 derived mutant peptide MSI 594F5A in lipopolysaccharide micelles, role of the helical hairpin conformation in outer-membrane permeabilization. J Am Chem Soc 132:18417–18428
Epand RM, Vogel HJ (1999) Diversity of antimicrobial peptides and their mechanisms of action. Biochim Biophys Acta 1462:11–28
Matsuzaki K (1998) Magainins as paradigm for the mode of action of pore forming polypeptides. Biochim Biophys Acta 1376:391–400
Shai Y (1999) Mechanism of the binding, insertion and destabilization of phospholipid bilayer membranes by α-helical antimicrobial and cell non-selective membrane-lytic peptides. Biochem Biophys Acta 1462:55–70
Bhunia A, Domadia PN, Torres J, Hallock KJ, Ramamoorthy A, Bhattacharjya S (2010) NMR structure of pardaxin, a pore-forming antimicrobial peptide, in lipopolysaccharide micelle mechanism of outer membrane permeabilization. J Biol Chem 285:3883–3895
Bhunia A, Mohanran H, Domadia PN, Torres J, Bhattacharjya S (2009) Designed b-boomerang antiendotoxic and antimicrobial peptides structures and activities in lipopolysaccharide. J Biol Chem 284:21991–22004
Raetz CR, Whitfield C (2002) Lipopolysaccharide endotoxins. Annu Rev Biochem 71:635–700
Rietschhel ET, Kirikae T, Schde FU, Mamat U, Schidt G, Loppnow H, Ulmer AJ, Zahringer U, Seydel U, DiPadova F, Schreuer M, Brade H (1994) Bacterial endotoxin: molecular relationships of structure to activity and function. FASEB J 8:217–225
Lad MD, Biirembaut F, Clifton LA, Frazier RA, Webster JRP (2007) Antimicrobial peptide-lipid binding interactions and binding selectivity. Biophys J 92:3575–3586
Delcour AH (2009) Outer membrane permeability and antibiotic resistance. Biochim Biophys Acta 1794:808–816
Hancock REW (1984) Alterations in outer membrane permeability. Annu Rev Microbiol 38:237–264
Hancock REW, Chapple DS (1999) Peptide antibiotics. Antimicrob Agents Chemother 43:1317–1323
Ding L, Yang L, Weiss TM, Warning AJ, Lehrer RI, Huang HW (2003) Interaction of antimicrobial peptides with lipopolysaccharides. Biochemistry 42:12251–12259
Bhattacharjya S (2010) De novo designed lipopolysaccharide binding peptides: structure based development of antiendotoxic and antimicrobial drugs. Curr Med Chem 17:3080–3093
Bringezu F, Wen S, Dante S, Hauss T, Majerowicz M, Waring A (2007) The insertion of the antimicrobial peptide dicynthaurin monomer in model membranes: thermodynamic and structural characterization. Biochemistry 46:5678–5686
Bruschi M, Pirri G, Giuliani A, Nicoletto SF, Baster I, Scorciapino MS, Casu M, Rinaldi AC (2010) Synthesis, characterization, antimicrobial activity and LPS-interaction properties of SB041, a novel dendrimeric peptide with antimicrobial properties. Peptides 31:1459–1467
Epand RM, Epand RF (2011) Bacterial membrane lipids in the action of antimicrobial agents. J Pept Sci 17:298–305
Hammer MU, Brauser A, Olak C, Brezesinski G, Goldmann T, Gutsmann T, Andra J (2010) Lipopolysaccharide interaction is decisive for the activity of the antimicrobial peptide NK-2 against Escherichia coli and Proteus mirabilis. Biochem J 427:477–488
Hartmann M, Berditsch M, Hawecker J, Ardakani MF, Gerthsen D, Ulrich AS (2010) Damage of the bacterial cell envelope by antimicrobial peptides Gramicidin S and PGLa as revealed by transmission and scanning electron microscopy. Antimicrob Agents Chemother 54:3132–3142
Junkes C, Harvey RD, Bruce KD, Dolling R, Bagheri M, Dathe M (2011) Cyclic antimicrobial R-, W-rich peptides: the role of peptide structure and E. coli outer and inner membranes in activity and mode of action. Eur Biophys J 40:515–528
Matsuzaki K, Sugishita K-I, Miyajima K (1999) Interactions of an antimicrobial peptide, magainin 2 with lipopolysaccharide-containing liposomes as a model for outer membranes of Gram-negative bacteria. FEBS Lett 449:221–224
Russell AL, Kennedy AM, Spuches A, Venugopal D, Bhonsle JB, Hicks RP (2010) Spectroscopic and thermodynamic evidence for antimicrobial peptide membrane selectivity. Chem Phys Lipids 163:488–497
Singh S, Kasetty G, Schmidtchen A, Malmsten M (2012) Membrane and lipopolysaccharide interactions of C-terminal peptides from S1 peptidases. Biochim Biophys Acta 18:2244–2251
Wieprecht T, Apostolov O, Seelig J (2000) Binding of the antibacterial peptide magainin 2 amide to small and large unilamellar vesicles. Biophys Chem 85:187–198
Ladokhin AS, Vidal MF, White SH (2010) CD spectroscopy of peptides and proteins bound to large unilamellar vesicles. J Membr Biol 236:247–253
Glattli A, Daura X, Seebach D, van Gunsteren WF (2002) Can one derive the conformational preference of a β-peptide from its CD spectrum. J Am Chem Soc 124:12972–12978
Ladokhin AS, Selsted ME, White SH (1999) CD spectra of indolicidin antimicrobial peptides suggest turns, not polyproline helix. Biochemistry 38:12313–12319
Wei S-T (2006) Solution structure of a novel tryptophan-rich peptide with bidirectional antimicrobial activity. J Bacteriol 188:328–334
Le Guemeve C, Auger M (1995) New approach to study fast and slow motions in lipid bilayers: application to dimyristoylphosphatidylcholine-cholesterol interactions. Biophys J 68:1952–1959
Andra J, Koch MH, Bartels R, Brandenburg K (2004) Antimicrob Agents Chemother 48:1593–1599
Brandenburg K, Kusumoto S, Seyed U (1997) Conformational studies of synthetic lipid A analogues and partial structures by infrared spectroscopy. Biochim Biophys Acta 2:183–201
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Hicks, R. (2017). Preparation of Membrane Models of Gram-Negative Bacteria and Their Interaction with Antimicrobial Peptides Studied by CD and NMR. In: Hansen, P. (eds) Antimicrobial Peptides. Methods in Molecular Biology, vol 1548. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6737-7_16
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DOI: https://doi.org/10.1007/978-1-4939-6737-7_16
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