Multiple Peptide Resistance Factor (MprF)-mediated Resistance of Staphylococcus aureus against Antimicrobial Peptides Coincides with a Modulated Peptide Interaction with Artificial Membranes Comprising Lysyl-Phosphatidylglycerol

Abstract

Modification of the membrane lipid phosphatidylglycerol (PG) of Staphylococcus aureus by enzymatic transfer of a L-lysine residue leading to lysyl-PG converts the net charge of PG from ؊1 to ؉1 and is thought to confer resistance to cationic antimicrobial peptides (AMPs). Lysyl-PG synthesis and translocation to the outer leaflet of the bacterial membrane are achieved by the membrane protein MprF. Consequently, mutants lacking a functional mprF gene are in particular vulnerable to the action of AMPs. Hence, we aim at elucidating whether and to which extent lysyl-PG modulates membrane binding, insertion, and permeabilization by various AMPs. Lysyl-PG was incorporated into artificial lipid bilayers, mimicking the cytoplasmic membrane of S. aureus. Moreover, we determined the activity of the peptides against a clinical isolate of S. aureus strain SA113 and two mutants lacking a functional mprF gene and visualized peptide-induced ultrastructural changes of bacteria by transmission electron microscopy. The studied peptides were: (i) NK-2, an ␣-helical fragment of mammalian NK-lysin, (ii) arenicin-1, a lugworm ␤-sheet peptide, and (iii) bee venom melittin. Biophysical data obtained by FRET spectroscopy, Fourier transform infrared spectroscopy, and electrical measurements with planar lipid bilayers were correlated with the biological activities of the peptides. They strongly support the hypothesis that peptide-membrane interactions are a prerequisite for eradication of S. aureus. However, degree and mode of modulation of membrane properties such as fluidity, capacitance, and conductivity were unique for each of the peptides. Altogether, our data support and underline the significance of lysyl-PG for S. aureus resistance to AMPs.

Further Insights into the Mode of Action of the Lipoglycopeptide Telavancin through Global Gene Expression Studies

“…K ϩ is the major cell cation, and increased production of lysine with its net positive charge may be a response to K ϩ depletion. Lysine is also a component of the positively charged phospholipid lysyl phosphatidylglycerol, and increases in its levels have been associated with increased surface positive charge and decreased susceptibility to multiple antimicrobial peptides (38) and with changes in membrane physical structure (1).…”

Low pH Enhances the Action of Maximin H5 against Staphylococcus aureus and Helps Mediate Lysylated Phosphatidylglycerol-Induced Resistance

“…It is generally accepted that the interaction with the CM of target bacteria is an essential step in the antimicrobial action of AMPs [13][14][15][16][17] and interactions of this type have been investigated using a variety of model membranes [18][19][20][21][22][23] . These model membranes are usually formed from lipid mixtures with a head-group composition that reflects the molar ratio of the major lipids in the bacterial membrane, namely PE, phosphatidylglycerol (PG), Lys-PG and cardiolipin (CL).…”

“…According to this mechanism of resistance, cationic AMPs activate a sensor histidine kinase (ApsS) of the ApsSR (GraSR) regulon, which leads to the up-regulated expression of the virulence factor mprF. The protein product of this gene, mprF, is a Lys-PG synthase that catalyses the transfer of lysine from lysyl-tRNA to the PG head-group and flipping of the resulting Lys-PG to the outer leaflet of the S. aureus CM 23,[31][32][33][34] . The presence of the lipid in the CM of S.…”

The lipid network

“…Furthermore, lipid modification in bacteria changes the effect of membraneactive molecules. For instance, the aminoacylation of PG and teichoic acid lipids in the Gram-positive S. aureus is indicated as a potential cause of AMP resistance because it increases the positive charges on the bacterial membrane surface, to ultimately "hide" the electrostatic attraction with cationic peptides (Andra et al 2011;Peschel 2002). Similar modification of LPS and lipid A were also observed in Gram-negative bacteria, where the addition of the amino acyl group reduced the binding of cationic molecules (Koprivnjak and Peschel 2011).…”