ISSN:
1365-2958
Source:
Blackwell Publishing Journal Backfiles 1879-2005
Topics:
Biology
,
Medicine
Notes:
Staphylococcal α-toxin forms heptameric pores that render membranes permeable for monovalent cations. The pore is formed by an amphipathic β-barrel encompassing amino acid residues 118–140 of each subunit of the oligomer. Human fibroblasts are susceptible to α-toxin but are able to repair the membrane lesions. Thereby, toxin oligomers remain embedded in the plasma membrane and exposed to the extracellular medium. In this study, we sought to detect structural changes occurring in the pore-forming sequence during lesion repair. Single cysteine substitution mutants were labelled with the environmentally sensitive fluorochrome acrylodan and, after mixing with wild-type toxin, incorporated into hybrid heptamers on fibroblast membranes. Formation of the lipid-inserted β-barrel was accompanied by characteristic fluorescence emission shifts. After lesion repair, the environment of the residues at the outer surface of the β-barrel remained unchanged, indicating continued contact with lipids. However, the labelled residues oriented towards the channel lumen underwent a green to blue shift in fluorescence, indicating reduced exposure to water. Pore closure proceeded in the presence of calmodulin inhibitors and of microtubule disruptors; however, it was prevented by cytochalasin D and by inhibitors of lipid metabolism. Our findings reveal the existence of a novel mechanism of membrane repair that may consist in constriction of the inserted proteinaceous pore within the lipid bilayer.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1046/j.1365-2958.2000.01865.x
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