Surface properties of membrane systems. Transport of staphylococcal δ-toxin from aqueous to membrane phase

doi:10.1016/0005-2736(77)90087-6

Biochimica et Biophysica Acta (BBA) - Biomembranes

Volume 465, Issue 2, 1 March 1977, Pages 378-390

https://doi.org/10.1016/0005-2736(77)90087-6 Get rights and content

Abstract

Hemolytic δ-toxin from Staphylococcus aureus was soluble in either water, methanol or chloroform/methanol (2 : 1, v/v). The toxin spread readily from distilled water into films with pressures (π) of 10 dynes/cm on water and 30 dynes/cm on 6 M urea; from chloroform/methanol it produced 40 dynes/cm pressure on distilled water. The toxin adsorbed barely from water ( $π = 1 dyne/cm$ ) but it did rapidly from 6 M urea ( $π = 35 dynes/cm$ ). The protein films had unusually high surface potentials, which increased with the film pressure and decreased with increasing both pH and urea concentration in the aqueous phase. The fluorescence of 1-aniline 8-naphthalene sulfonate with δ-toxin was much greater than that with RNAase and dipalmitoyl phosphatidylcholine itself, indicating probably a marked lipid-binding character of the toxin. By circular dichroism the α-helix content of δ-toxin was 42% in water, 45% in methanol, 24% in 6 M urea. Infrared spectroscopy showed predominant α-helix in both ²H₂O and deuterated chloroform/methanol as well as in films spread from either solvent on ²H₂O. In spreading from 6 M [²H]urea, in which the major infrared absorption was that of [²H]urea with peaks at 1600 and 1480 cm⁻¹, and δ-toxin film showed prevalently non-α-helix structures with major peak intensities at 1633 cm⁻¹ > 1680 cm⁻¹, indicating the appearance of new β-aggregated and β-antiparallel pleated sheet structures in the film. The data prove that (1) high pressure protein films can consist of α-helix as well as non-α-helix structures and, differently from another cytolytic protein, melittin, δ-toxin does not resume the α-helix conformation in going into the film phase from the extended chain in 6 M urea; (2) conformational changes are important in the transport of proteins from aqueous to lipid or membrane phase; (3) δ-toxin is by far more versatile in structural dynamics and more surface active than α-toxin.

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Staphylococcus aureus δ-toxin in aqueous solution: Behavior in monomeric and multimeric states
2017, Biophysical Chemistry
Citation Excerpt :
In fact, previous MD simulation studies have explored and validated such aspects, as evaluating δ-toxin interaction with POPC lipid bilayers in atomic-level [18], and coarse-grained resolutions [19]. In aqueous solution, however, it adopts varying helical content, which is dependent on its concentration [20,21,11,22]. Although this much knowledge about δ-toxin behavior, there is no structural, atomic-level data available for this peptide in aqueous solutions.
δ-Toxin is a 26 amino acid peptide capable of lysing several mammalian cell types and subcellular structures. Structurally, δ-toxin predominantly exhibits a α-helical secondary structure in membranes but, in aqueous solution, it adopts varying helical content. As no atomic-level data is available for this peptide in aqueous solutions and for the water-to-membrane transition, this work aims to characterize δ-toxin behavior in these conditions through molecular dynamics simulations in triplicates employing four different parameter sets. Our results, validated on previous experimental data, suggest that the peptide has from 4 to 16 residues folded as α-helix in aqueous solution, and a water-to-membrane foldamer comprising residues 14–18. Considering a previously proposed stable tetramer form in aqueous solutions, protein-protein docking and molecular dynamics studies were performed, suggesting that δ-toxin increases its α-helical content in such organization. The obtained results are expected to contribute in future studies on δ-toxin aggregation and interaction to biomembranes.
Purification and characterization of a novel delta-lysin variant that inhibits Staphylococcus aureus and has limited hemolytic activity
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Delta-lysins (DL) that are produced by various species of staphylococci are not widely known for their antimicrobial activity. We have purified and characterized a novel DL variant, E229DL and examined its spectrum of inhibitory activity. The biological activity of E229DL, produced by Staphylococcus epidermidis strain E229, shows relatively broad-spectrum activity against Gram-positive pathogens, including representatives of MRSA and epidemic MRSA type 15. E229DL was purified to homogeneity from 95% acidified-methanol extracts of cell cultures by using a series of reversed-phase chromatographic separations. The fully processed form of E229DL is a 25-amino-acid peptide with a predicted mass of 2841.4 Da, but the purified biologically active molecule appears to be N-formylated (mass 2867.33 Da). The DL gene (hld) resembles that of other types of DL, but differs in five codons with hld in Staphylococcus aureus (26 residues) and one codon with the closest homolog, the hld-II in S. warneri (25 residues). The characterization of E229DL showed that its activity is stable in agar exposed to high temperatures (80 °C/45 min). In addition, biological testing of the native and synthetic peptides against a range of human and animal erythrocytes and Vero cells indicated that E229DL is an antibacterial agent with no detectable cytopathic effects at concentrations equivalent to the minimum inhibitory concentration for EMRSA15-A208. Initial investigation of the mode of action of E229DL indicated that it is rapidly lytic for target cells. This is the first description of a native form of DL having only limited cytotoxic activity for eukaryotic cells at concentrations that are inhibitory to staphylococci.
δ-hemolysin, an update on a membrane-interacting peptide
2009, Peptides
δ-hemolysin is a hemolytic peptide produced by Staphylococcus, and it has been studied for nearly 50 years. Therefore, it has become a model in the study of peptides interacting with membranes. In this review, we report some recent findings and compare them with previous works. δ-hemolysin is a 26 amino acid peptide, somewhat hydrophobic and presenting a zero net charge. Study of its structure has shown that δ-hemolysin is α-helical and amphipathic, such as many antimicrobial peptides (e.g. magainin and melittin). However, δ-hemolysin had not displayed any reported antimicrobial activity until a recent publication showed its high potency against Legionella. Its mode of action is based on direct interaction with target membranes. In accordance with its concentration, δ-hemolysin may slightly perturb a membrane or lead to cell lysis. Peptide charge plays an important role in its interaction with membranes, as is shown in the study of peptide variants. Some positively charged variants become highly hemolytic and even active against Escherichia coli and Staphylococcus aureus. Finally, it has recently been demonstrated that peptide preferentially binds to lipid-disordered domains. It has been postulated that as a result, enrichment in lipid-ordered domains might increase peptide concentration in lipid-disordered domains and thereby improve its activity.
Molecular dynamics simulations of the insertion of two ideally amphipathic lytic peptides LK<inf>15</inf> and LK<inf>9</inf> in a 1,2-dimyristoylphosphatidylcholine monolayer
2001, Biochimica et Biophysica Acta - Biomembranes
Citation Excerpt :
β-Sheet structures are also involved but with very different peptides [3,1]. In order to reproduce the behavior of natural amphipathic peptidic toxins like melittin [4], δ-hemolysin, bombolittin and defensin, artificial toxins designed with a minimalist approach were synthesized: only two amino-acids were used, the most lipophilic (L) and the most hydrophilic (K), also with a 2:1 L/K ratio, these peptides are designed to have a single charged K residue per putative α-helical turn to generate ideally amphipathic helices. Like natural peptides, the LiKj (i=2j) peptides are surface active because of their secondary amphipathic character and they have been shown to be more efficient than natural peptides [5].
We present here the results of 1-ns molecular dynamics (MD) simulations of two ideally amphipathic lytic peptides, namely LK₁₅ and LK₉, in a 1,2-dimyristoylphosphatidylcholine monolayer with two different cross-sectional areas per lipid of 80 Å² (loose film) and 63 Å² (tight standard film). These peptides are lytic, ideally amphipathic with a minimalist composition L_iK_j and the following sequences: H₂N-KLLKLLLKLLLKLLK-CO-Ph for LK₁₅ and H₂N-KLKLKLKLK-CO-Ph for LK₉. From experimental data, LK₁₅ exhibits an α-helical secondary structure, whereas LK₉ was found to form antiparallel β-sheets at the interface of a DMPC monolayer. Whatever the specific lipid surface is, the two peptides exhibit very different behavior: the α-helix inserts deeply into the monolayer whereas the β-sheeted peptide stays at the surface within the upper polar part of the monolayer. In all cases, a loose monolayer (80 Å²) results in noticeable artifacts whereas a monolayer with standard specific surface leads to very reliable behavior well in accordance with experimental data. Despite their different insertion depth, the two peptides exhibit identical lytic efficiency. This is very likely a direct consequence of the very strong Van der Waals interactions between the fatty alkyl chains of the lipids and the highly lipophilic lower part of the peptide, resulting in an identical thinning of the two monolayers.
The monolayer technique: A potent tool for studying the interfacial properties of antimicrobial and membrane-lytic peptides and their interactions with lipid membranes
1999, Biochimica et Biophysica Acta - Biomembranes
Erudites of the antiquity already knew the calming effect of oil films on the sea waves. But one had to wait until 1774 to read the first scientific report on oil films from B. Franklin and again 1878 to learn the thermodynamic analysis on adsorption developed by J. Gibbs. Then, in 1891, Agnes Pockels described a technique to manipulate oil films by using barriers. Finally, in 1917, I. Langmuir introduced the experimental and theoretical modern concepts on insoluble monolayers. Since that time, and because it has been found to provide invaluable information at the molecular scale, the monolayer technique has been more and more extensively used, and, during the past decade, an explosive increase in the number of publications has occurred. Over the same period, considerable and ever-increasing interest in the antimicrobial peptides of various plants, bacteria, insects, amphibians and mammals has grown. Because many of these antimicrobial peptides act at the cell membrane level, the monolayer technique is entirely suitable for studying their physicochemical and biological properties. This review describes monolayer experiments performed with some of these antimicrobial peptides, especially gramicidin A, melittin, cardiotoxins and defensin A. After giving a few basic notions of surface chemistry, the surface-active properties of these peptides and their behavior when they are arranged in monomolecular films are reported and discussed in relation to their tridimensional structure and their amphipathic character. The penetration of these antimicrobial peptides into phospholipid monolayer model membranes, as well as their interactions with lipids in mixed films, are also emphasized.
Possible virulence factors of Staphylococcus aureus in a mouse septic model
1999, FEMS Immunology and Medical Microbiology
Twenty clinical isolates of Staphylococcus aureus were examined to elucidate the virulence factors which are directly related to lethality in a mouse septic model. Heat or formalin treatment of the organism abolished the lethal activity of the live organism during challenge intravenously administered via the tail vein. Nevertheless, injection of ten times concentrated culture supernatant fluid (SUP) showed lethal activity in the mouse. However, there was no lethality when SUP was heated at 60°C for 15 min. To examine variations of SUP lethality among strains, we collected 20 strains of S. aureus from four different hospitals. Then, we compared several factors for SUP lethality, which were the extracellular toxins and enzymes, such as toxic shock syndrome toxin 1, enterotoxin A, B, D, and hemolysins (α,β,γ), and also cytotoxic activity to human polymorphonuclear leukocytes and Vero cells. No difference was found among these factors except cytotoxic activity to Vero cells. Furthermore, we compared two strains in a mouse septic model according to the grade of bacteremia and lethal events. We found that mortality was higher with challenge by the strain whose SUP was lethal in comparison to the strain whose SUP was not lethal, even though the viable bacteria counts in the septic blood in both strains were not significantly different. This strongly supports the possibility that extracellular products, not the cell wall components, of S. aureus play the key role in the lethal event in this mouse septic model. In addition, among the extracellular products, those which have cytotoxic activity to Vero cells may contribute to the lethality in sepsis caused by S. aureus in this murine model.

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Surface properties of membrane systems. Transport of staphylococcal δ-toxin from aqueous to membrane phase

Abstract

Biochim. Biophys. Acta

J. Biol. Chem.

J. Colloid Interface Sci.

Biochim. Biophys. Acta

Arch. Biochem. Biophys.

J. Lipid Res.

Biochim. Biophys. Acta

Biochim. Biophys. Acta

Chem. Phys. Lipids

Biochim. Biophys. Acta

J. Colloid Interface Sci.

Biochim. Biophys. Acta

Lipids

Ann. N.Y. Acad. Sci.

Science

J. Bacteriol.