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Characterization of a Williopsis saturnus var. mrakii high molecular weight secreted killer toxin with broad-spectrum
Cyril Guyard1, Nathalie Séguy, Jean-Charles Cailliez
1Département de Microbiologie des Ecosystèmes, IFR17, Institut Pasteur de Lille, 1 rue Calmette, BP 245, 59019 Lille, France. Cyril.Guyard@pasteur-lille.fr
Abstract:
Williopsis saturnus var. mrakii MUCL 41968 secretes a killer toxin (WmKT), which is active against a wide range of pathogens. From the W. saturnus var. mrakii MUCL 41968 culture supernatant a protein of 85 kDa with killer activity was purified to homogeneity. The purified protein was demonstrated to be a killer toxin since it displays the toxin activity and cross-reacts with mAbKT4, a monoclonal antibody that blocks WmKT activity. Its partial amino acid sequencing revealed that WmKT might be related to yeast SUN proteins, but not to other killer toxins described. Immunofluorescence studies using polyclonal antibodies raised against purified WmKT revealed that it acts by binding to the cell surface of sensitive strains. We showed that WmKT is inactive against mutant strains of Saccharomyces cerevisiae deficient in the synthesis of beta-glucans, indicating that these polysaccharides constitute the target of the toxin. WmKT was demonstrated to induce rapid lethal cell permeation, since strong propidium iodide labelling was shown for sensitive strains treated with the killer toxin. These findings indicate that WmKT is a novel killer toxin whose molecular characterization may lead to the development of new wide-spectrum antimicrobial compounds.
Insights
A novel yeast killer toxin, Williopsis saturnus var. mrakii killer toxin (WmKT), was purified and characterized. This toxin targets beta-glucans on cell surfaces, causing lethal permeation and showing potential for new antimicrobial development.
Area of Science:
- Microbiology
- Biochemistry
- Yeast Genetics
Background:
- Williopsis saturnus var. mrakii produces a killer toxin (WmKT) effective against various pathogens.
- Killer toxins are proteins secreted by yeasts that inhibit or kill other sensitive yeast strains.
- Understanding novel killer toxins can lead to new antimicrobial strategies.
Purpose of the Study:
- To purify and characterize the killer toxin secreted by W. saturnus var. mrakii MUCL 41968.
- To elucidate the molecular mechanism and target of the killer toxin.
- To assess the potential of the toxin as a novel antimicrobial agent.
Main Methods:
- Purification of the 85 kDa protein from culture supernatant.
- Confirmation of killer activity and cross-reactivity with a specific monoclonal antibody (mAbKT4).
- Partial amino acid sequencing, immunofluorescence, and testing against mutant Saccharomyces cerevisiae strains deficient in beta-glucan synthesis.
- Propidium iodide staining to assess cell permeation.
Main Results:
- An 85 kDa protein with killer activity was purified and identified as WmKT.
- WmKT showed relation to yeast SUN proteins, distinct from other known killer toxins.
- The toxin binds to the cell surface of sensitive strains.
- WmKT is inactive against beta-glucan-deficient yeast mutants, identifying beta-glucans as the target.
- WmKT induces rapid cell permeation, leading to cell death.
Conclusions:
- Williopsis saturnus var. mrakii killer toxin (WmKT) is a novel toxin with a unique mechanism of action.
- Beta-glucans are the specific target of WmKT on the cell surface.
- The molecular characterization of WmKT opens avenues for developing new broad-spectrum antimicrobial compounds.
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