1. Fluorescence studies of bacterial membrane proteins and cell signalling
- Author
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Fišer, Radovan, Konopásek, Ivo, Hof, Martin, and Forstová, Jitka
- Subjects
Fura-2 ,makrofág ,planární lipidové membrány ,membránový kanál ,vápníková signalizace ,endocytóza ,translokace proteinu ,endocytosis ,narušování lipozómů ,liposome disruption ,membrane channels ,CyaA ,Bordetella ,calcium signalling ,protein translocation ,planar lipid membranes ,macrophage - Abstract
(English) This work is based on five publications studying mostly adenylate cyclase toxin (CyaA) from Bordetella pertussis and its interaction with biological membranes. CyaA permeabilizes cell membranes by forming small cationselective pores and subverts cellular signaling by delivering an adenylate cyclase (AC) enzyme that converts ATP to cAMP into host cells. First study clarifies the membrane disruption mechanisms of CyaA and another bacterial RTX toxin; αhemolysin (HlyA) from Escherichia coli. For this purpose, we employed a fluorescence requenching method using liposomes as target membranes. We showed that both toxins induced a graded leakage of liposome content with different ion selectivities (Fišer a Konopásek 2009). Both AC delivery and pore formation were previously shown to involve a predicted amphipathic αhelix(502522). In the second publication we investigated another predicted transmembrane αhelix(565591) that comprises a Glu(570) and Glu(581) pair. We examined the roles of these glutamates in the activity of CyaA, mostly on planar lipid membranes end erythrocytes. Negative charge at position 570, but not at position 581, was found to be essential for cation selectivity of the pore, suggesting a role of Glu(570) in...
- Published
- 2011