Lactoferricin B causes depolarization of the cytoplasmic membrane of <i>Escherichia coli</i> ATCC 25922 and fusion of negatively charged liposomes

Ulvatne, Hilde; Haukland, Hanne H.; Olsvik, Ørjan; Vorland, Lars H.

doi:10.1016/s0014-5793(01)02233-5

Cited by 82 publications

(67 citation statements)

References 22 publications

(21 reference statements)

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“…The disulfide cross linked, antimicrobial β-hairpin peptide lactoferricin retains its antimicrobial biological activity even after 19 of its 25 residues, including all of the disulfide cross links are removed, requiring only a core linear hexapeptide that is as active as the 25-residue disulfide-cross linked parent β-hairpin (55). In fact many derivatives of lactoferricin are highly active antimicrobial peptides (56)(57)(58). Disulfide cross linked variants of our FSKRGY have antimicrobial activity that is indistinguishable from the non-cross linked peptide (JRM and WCW, unpublished observations).…”

Section: Correlation Between Structure and Function In Membranesmentioning

confidence: 99%

β-Sheet Pore-Forming Peptides Selected from a Rational Combinatorial Library: Mechanism of Pore Formation in Lipid Vesicles and Activity in Biological Membranes

et al. 2007

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In a previous report we described the selection of potent, β-sheet pore-forming peptides from a combinatorial library designed to mimic membrane-spanning β-hairpins (Rausch JM, Marks JR and Wimley WC, (2005) PNAS, 102:10511-5). Here, we characterize their mechanism of action and compare the structure-function relationships in lipid vesicles to their activity in biological membranes. The pore-forming peptides bind to membrane interfaces and self-assemble into β-sheets that cause a transient burst of graded leakage across the bilayers. Despite the continued presence of the structured peptides in the bilayer, at most peptide concentrations leakage is incomplete and ceases quickly after peptide addition with a deactivation half-time of several minutes. Molecules up to 3,000 Da escape from the transient pores, but much larger molecules do not. Fluorescence spectroscopy and quenching showed that the peptides reside mainly on the bilayer surface and are partially exposed to water, rather than in a membrane-spanning state. The "carpet" or "sinking raft" model of peptide pore formation offers a viable explanation for our observations and suggests that the selected pore formers function with a mechanism that is similar to the natural pore-forming antimicrobial peptides. We therefore also characterized the antimicrobial and cytotoxic activity of these peptides. All peptides studied, including non pore-formers, had sterilizing antimicrobial activity against at least some microbes, and most have low activity against mammalian cell membranes. Thus, the structurefunction relationships that were apparent in the vesicle systems are similar to, but do not correlate completely with the activity of the same peptides in biological membranes. However, of the peptides tested, only the pore-formers selected in the high throughput screen have potent, broad-spectrum sterilizing activity against Gram-positive and Gram-negative bacteria as well as against fungi, while having only small lytic effects on human cells.A wide variety of pore-forming proteins and peptides take advantage of the discontinuous hydrophobicity of membrane-spanning β-sheets, which makes them well suited for undergoing transitions between hydrophilic and hydrophobic environments. For example, the β-barrel protein toxins including perfringolysin-O (1), α-hemolysin (2), and the anthrax protective antigen(3) pre-assemble a protein scaffold on membrane surfaces and then undergo a spontaneous transition in which loop sequences change conformation and are inserted into the membrane to form β-barrel pores. Similarly, there are many pore-forming, antimicrobial peptides (AMPs) 1 that are soluble in water and which self assemble into β-sheet-rich pores in membranes(4-6). To explore the structural determinants of folding and self-assembly of β- † Supported by the National Institutes of Health grant GM060000 *Corresponding author: Phone: 504-988-7076; Fax:504-988-2739, Email: wwimley@tulane.edu. NIH Public Access Author ManuscriptBiochemistry. Author manuscript; available ...

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Section: Correlation Between Structure and Function In Membranesmentioning

confidence: 99%

β-Sheet Pore-Forming Peptides Selected from a Rational Combinatorial Library: Mechanism of Pore Formation in Lipid Vesicles and Activity in Biological Membranes

et al. 2007

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“…The bactericidal effect of many CAPs is thought to be due to the action on the cytoplasmic membrane of the susceptible bacteria, possibly through the formation of pores or destabilization of the membrane bilayer structure leading to lysis of the cell [4,5]. Some CAPs such as buforin 11 [6], indolicidin [7], apidaecin [8], PR 39 [9], PR 26 [10], bactenecin [11] and lactoferricin B [12,13] have been reported to have intracellular targets. In a recent review article, Wimley [14] pointed out that many antimicrobial peptides do not porate or damage bacterial membranes but rather produce transient pores as demonstrated by using unilamellar liposomes loaded with probes.…”

Section: Introductionmentioning

confidence: 99%

Interaction of protamine with gram‐negative bacteria membranes: possible alternative mechanisms of internalization in Escherichia coli, Salmonella typhimurium and Pseudomonas aeruginosa

Pink

Hasan

Quinn

et al. 2014

Journal of Peptide Science

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This study was concerned with the interaction between the cationic antimicrobial peptide, protamine (Ptm) and the cytoplasmic membranes of the gram-negative bacteria Escherichia coli, Salmonella typhimurium and Pseudomonas aeruginosa. The objective of the study was to explain the observed paradox of internalization without permanent disruption of the cell envelope. We carried out Monte Carlo computer simulation of Ptm in an aqueous environment in the presence of~100 mM NaCl and model membranes consisting of either (65:35) or (75:25) PE:PG molar ratios. The (75:25) model, representative of the gram-negative cytoplasmic membrane, showed that the Ptm center of mass remained at least 7 nm from the membrane surface leading to the prediction that Ptm would not internalize via disruption of the inner membrane.By using immunoelectron microscopy of Ptm-treated cells, we showed that Ptm internalization to the cytoplasm took place rapidly in the presence or absence of the outer envelope. Ultrastructural examination revealed no obvious morphological changes to cells that were treated with subinhibitory or bactericidal levels of Ptm. Reconstituted phospholipid bilayers were constructed and were unperturbed by Ptm treatment over a wide range of concentrations and applied transmembrane voltages. We conclude that in the cases of the cell envelopes of E. coli, S. typhimurium and P. aeruginosa, Ptm internalized by means independent of the phospholipid bilayer, most likely mediated by one or more membrane proteins such as cation-selective barrel-like proteins. Work is currently underway to test this hypothesis.

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“…Some AMPs have been proven to penetrate the cell membrane and affect the intracellular targets within the cells (11)(12)(13); Lfcin B is one of these AMPs (14 -18). Some studies have indicated that Lfcin B leads to the depolarization of the cell membrane and does not lyse the cells (19); Lfcin B has been shown to inhibit the macromolecular synthesis of cells, which suggests that the intracellular targets of Lfcin B may exist (12,19,20).…”

mentioning

confidence: 99%

Lactoferricin B Inhibits the Phosphorylation of the Two-Component System Response Regulators BasR and CreB

Sung

Chen

2012

Molecular & Cellular Proteomics

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Natural antimicrobial peptides provide fundamental protection for multicellular organisms from microbes, such as Lactoferricin B (Lfcin B). Many studies have shown that Lfcin B penetrates the cell membrane and has intracellular activities. To elucidate the intracellular behavior of Lfcin B, we first used Escherichia coli K12 proteome chips to identify the intracellular targets of Lfcin B. The results showed that Lfcin B binds to two response regulators, BasR and CreB, of the two-component system. For further analysis, we conducted several in vitro and in vivo experiments and utilized bioinformatics methods. The electrophoretic mobility shift assays and kinase assays indicate that Lfcin B inhibits the phosphorylation of the response regulators (BasR and CreB) and their cognate sensor kinases (BasS and CreC). Antibacterial assays showed that Lfcin B reduced E. coli's tolerance to environmental stimuli, such as excessive ferric ions and minimal medium conditions. This is the first study to show that an antimicrobial peptide inhibits the growth of bacteria by influencing

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Lactoferricin B causes depolarization of the cytoplasmic membrane of Escherichia coli ATCC 25922 and fusion of negatively charged liposomes

Cited by 82 publications

References 22 publications

β-Sheet Pore-Forming Peptides Selected from a Rational Combinatorial Library: Mechanism of Pore Formation in Lipid Vesicles and Activity in Biological Membranes

β-Sheet Pore-Forming Peptides Selected from a Rational Combinatorial Library: Mechanism of Pore Formation in Lipid Vesicles and Activity in Biological Membranes

Interaction of protamine with gram‐negative bacteria membranes: possible alternative mechanisms of internalization in Escherichia coli, Salmonella typhimurium and Pseudomonas aeruginosa

Lactoferricin B Inhibits the Phosphorylation of the Two-Component System Response Regulators BasR and CreB

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