[en] alpha-Haemolysin is an extracellular protein toxin (approximately 107 kDa)
secreted by Escherichia coli that acts at the level of the plasma membranes of
target eukaryotic cells. The nature of the toxin interaction with the membrane is
not known at present, although it has been established that receptor-mediated
binding is not essential. In this work, we have studied the perturbation produced
by purified alpha-haemolysin on pure phosphatidylcholine bilayers in the form of
large unilamellar vesicles, under conditions in which the toxin has been shown to
induce vesicle leakage. The bilayer systems containing bound protein have been
examined by differential scanning calorimetry, fluorescence spectroscopy,
differential solubilization by Triton X-114, and freeze-fracture electron
microscopy. All the data concur in indicating that alpha-haemolysin, under
conditions leading to cell lysis, becomes inserted in the target membrane in the
way of intrinsic or integral proteins. In addition, the experimental results
support the idea that inserted alpha-haemolysin occupies only one of the membrane
phospholipid monolayers, i.e. it is not a transmembrane protein. The experimental
data are complemented by structure prediction studies according to which as many
as ten amphipathic alpha-helices, appropriate for protein-lipid interaction, but
no hydrophobic transmembrane helices are predicted in alpha-haemolysin. These
observations and predictions have important consequences for the mechanism of
cell lysis by alpha-haemolysin; in particular, a non-transmembrane arrangement of
the toxin in the target membrane is not compatible with the concept of
alpha-haemolysin as a pore-forming toxin.
Disciplines :
Biochemistry, biophysics & molecular biology
Author, co-author :
Soloaga, A.
Veiga, Mp.
Garcia-Segura, Lm.
Ostolaza, H.
Brasseur, Robert ; Université de Liège - ULiège > Gembloux Agro-Bio Tech
Goni, Fm.
Language :
English
Title :
Insertion Of Escherichia Coli Alpha-Haemolysin In Lipid Bilayers As A Non-Transmembrane Integral Protein: Prediction And Experiment
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