Related Experiment Videos
Interaction of eosinophil granule major basic protein with synthetic lipid bilayers: a mechanism for toxicity
R I Abu-Ghazaleh1, G J Gleich, F G Prendergast
1Department of Biochemistry and Molecular Biology, Mayo Clinic, Rochester, Minnesota 55905.
Abstract:
Eosinophil granule major basic protein (MBP) is a potent toxin for mammalian cells and helminths, but the mechanism of its toxicity is not known. Here we tested whether MBP toxicity is exerted through its effect on the lipid bilayer of its targets. Liposomes prepared from synthetic phospholipids were used as targets for MBP and their properties examined by fluorescence and circular dichroism (CD) spectroscopy. MBP caused a change in the temperature transition profiles of acidic liposomes (1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidyl serine or an equimolar mixture of 1,2-dimyristoyl-sn-glycero-3-phosphocholine and 1,2-dimyristoyl-sn-glycero-3-phosphatidic acid) and induced their aggregation as shown by fluorescence resonance energy transfer experiments. The CD spectra and fluorescence characteristics of MBP itself were altered by its interaction with acidic lipids. Blue shifts in the emission maxima of the Trp, and of the dimethylaminonaphthyl moiety in acrylodan-labeled MBP, and a reduction in the effectiveness of quenching of Trp fluorescence by acrylamide were observed in the presence of acidic lipids. None of these effects were noted with zwitterionic lipids. This MBP: lipid bilayer interaction resulted in fusion and lysis of liposomes as indicated by the fluorescent indicator calcein. The results demonstrate that MBP associates with acidic lipids and that it disrupts, aggregates, fuses, and lyses liposomes prepared from such lipids. Such interaction might account for its wide range of toxicity.
Insights
Eosinophil major basic protein (MBP) targets acidic lipids, disrupting, fusing, and lysing liposomes. This interaction with lipid bilayers may explain MBP's broad toxicity to cells and helminths.
Area of Science:
- Biochemistry
- Cell Biology
- Toxicology
Background:
- Eosinophil granule major basic protein (MBP) is a potent toxin.
- The mechanism underlying MBP's toxicity remains largely unknown.
- Investigating MBP's interaction with target membranes is crucial for understanding its cytotoxic effects.
Purpose of the Study:
- To determine if MBP exerts its toxicity by affecting the lipid bilayer of target cells.
- To elucidate the molecular interactions between MBP and phospholipid bilayers.
- To understand how MBP binding leads to membrane disruption and cell lysis.
Main Methods:
- Liposomes composed of synthetic phospholipids were used as model targets.
- Fluorescence spectroscopy and circular dichroism (CD) spectroscopy were employed to analyze MBP-lipid interactions.
- Fluorescence resonance energy transfer (FRET) and calcein leakage assays were used to assess liposome aggregation, fusion, and lysis.
Main Results:
- MBP altered the temperature transition profiles and induced aggregation of acidic liposomes.
- MBP's interaction with acidic lipids caused changes in its own fluorescence and CD spectra.
- MBP induced liposome fusion and lysis, evidenced by calcein release, specifically with acidic phospholipids, not zwitterionic ones.
Conclusions:
- MBP directly associates with and disrupts acidic lipid bilayers.
- MBP induces liposome aggregation, fusion, and lysis through interaction with acidic phospholipids.
- These membrane-disrupting properties likely account for MBP's broad toxicity.