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Nano-viscosimetry analysis of the membrane disrupting action of the bee venom peptide melittin.

ABSTRACT: Melittin is one of the most studied ?-helical cationic membrane disrupting peptides. It is the main component of bee venom, however it is considered an antimicrobial peptide for its ability to kill bacteria. Melittin is believed to act by opening large toroidal pores in the plasma membrane of the targeted cells/bacteria, although this is questioned by some authors. Little is known, however, about the molecular mechanism leading to this activity. In this study the mechanism of action of melittin was studied by dye leakage and quartz crystal microbalance fingerprinting analysis in biomimetic model membranes. The results revealed the existence of multiple stages in the membrane disrupting action with characteristic differences between different membrane types. In bacterial-mimetic (charged) lipid mixtures the viscoelastic fingerprints suggest a surface-acting mechanism, whereas in mammalian-mimetic (neutral) membranes melittin appears to penetrate the bilayer already at low concentrations. In domain-forming mixed membranes melittin shows a preference for the domain containing predominantly zwitterionic lipids. The results confirm membrane poration but are inconsistent with the insertion-to-toroidal pore pathway. Therefore hypotheses of the two membrane disrupting pathways were developed, describing the membrane disruption as either surface tension modulation leading to toroidal pore formation, or linear aggregation leading to fissure formation in the membrane.

SUBMITTER: Pandidan S

PROVIDER: S-EPMC6658469 | biostudies-literature | 2019 Jul

REPOSITORIES: biostudies-literature

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Nano-viscosimetry analysis of the membrane disrupting action of the bee venom peptide melittin.

Pandidan Sara S Mechler Adam A

Scientific reports 20190725 1

Melittin is one of the most studied α-helical cationic membrane disrupting peptides. It is the main component of bee venom, however it is considered an antimicrobial peptide for its ability to kill bacteria. Melittin is believed to act by opening large toroidal pores in the plasma membrane of the targeted cells/bacteria, although this is questioned by some authors. Little is known, however, about the molecular mechanism leading to this activity. In this study the mechanism of action of melittin ...[more]

PMID: 31346251

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Project description:Melittin is a basic 26-amino-acid polypeptide that constitutes 40-60% of dry honeybee (Apis mellifera) venom. Although much is known about its strong surface activity on lipid membranes, less is known about its pain-producing effects in the nervous system. In this review, we provide lines of accumulating evidence to support the hypothesis that melittin is the major pain-producing substance of bee venom. At the psychophysical and behavioral levels, subcutaneous injection of melittin causes tonic pain sensation and pain-related behaviors in both humans and animals. At the cellular level, melittin activates primary nociceptor cells through direct and indirect effects. On one hand, melittin can selectively open thermal nociceptor transient receptor potential vanilloid receptor channels via phospholipase A2-lipoxygenase/cyclooxygenase metabolites, leading to depolarization of primary nociceptor cells. On the other hand, algogens and inflammatory/pro-inflammatory mediators released from the tissue matrix by melittin's pore-forming effects can activate primary nociceptor cells through both ligand-gated receptor channels and the G-protein-coupled receptor-mediated opening of transient receptor potential canonical channels. Moreover, subcutaneous melittin up-regulates Nav1.8 and Nav1.9 subunits, resulting in the enhancement of tetrodotoxin-resistant Na(+) currents and the generation of long-term action potential firing. These nociceptive responses in the periphery finally activate and sensitize the spinal dorsal horn pain-signaling neurons, resulting in spontaneous nociceptive paw flinches and pain hypersensitivity to thermal and mechanical stimuli. Taken together, it is concluded that melittin is the major pain-producing substance of bee venom, by which peripheral persistent pain and hyperalgesia (or allodynia), primary nociceptive neuronal sensitization, and CNS synaptic plasticity (or metaplasticity) can be readily induced and the molecular and cellular mechanisms underlying naturally-occurring venomous biotoxins can be experimentally unraveled.

Project description:The ability of the peptides melittin, [Ala-14]melittin (P14A) and whole bee venom to lyse red blood cells (RBC) and to cause shape transformation, binding, partitioning and changes in volume of the cells during haemolysis, as well as the action of the bivalent cations Zn2+ and Ca2+, chlorpromazine, albumin and plasma on the peptide-induced haemolysis of RBC in high ionic-strength solution, have been investigated. The protective effect of all inhibitors depends on whether they have been added to the media before or after the cells. When added before the cells they reduced significantly the rate of peptide-induced haemolysis and shape transformation. The effect was maximal when agents acted simultaneously after introduction of the cells into the media containing both inhibitors and peptides. Incubation of the cells in isotonic solution before the addition of peptides enhanced 2-3-fold the RBC susceptibility (i.e. rate of haemolysis) to lytic action of the same amount of peptides, and increased the order of the haemolytic reaction, although the power law coefficient did not exceed a value of 2 for all peptides, suggesting that haemolysis is attributable to the monomeric or dimeric forms of the peptides. Partition coefficients were of the order of approximately 10(6) M-1, and P14A possessed a value 3-fold larger compared with melittin and bee venom, which correlated with its enhanced haemolytic activity. The protective action of inhibitors against peptide-induced haemolysis has been explained on the basis of their ability to compete with peptide binding at an early stage of peptide-membrane interaction, and not as a result of inhibition of a pre-existing peptide-induced pore. Whereas melittin increased the volume of RBC during haemolysis, P14A, melittin in the presence of phospholipase A2 or bee venom, reduced the volume in a concentration-dependent manner. The present data reveal the significant role of the initial stage of peptide-membrane interaction and peptide structure in the mechanism of haemolysis. These data are not consistent with a lipid-based mechanism of peptide-induced haemolysis, indicating that the mode of peptide-protein interaction is an important and decisive step in the haemolytic mechanism. It should be noted that data (in the form of three additional Tables) on the ability of inhibitors to protect cells from haemolysis when inhibitor and peptide act simultaneously are available. They are reported in Supplementary Publication SUP 50178, which has been deposited at the British Library Document Supply Centre, Boston Spa, Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1996) 313, 9.

Dataset Information

Nano-viscosimetry analysis of the membrane disrupting action of the bee venom peptide melittin.

Publications

Nano-viscosimetry analysis of the membrane disrupting action of the bee venom peptide melittin.

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