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The role of a conserved tyrosine residue of acylated domain in membrane insertion and penetration of RTX toxins
Lepesheva, Anna ; Mašín, Jiří (advisor) ; Petráčková, Denisa (referee)
Pore-forming RTX toxins are key virulence factors of many Gram-negative pathogens. These proteins share several common structural and functional features: (i) the presence of repetitive sequences rich in glycine and aspartate, which are important for calcium ion binding, (ii) transport from the bacterial cytosol through the type I secretion system (T1SS), (iii) modification by a fatty acid at specific lysines in the acylated domain by a specific acyltransferase, and (iv) the presence of an amphipathic region responsible for the formation of cation-selective pores in the target membrane. The aromatic side chain of the conserved tyrosine residue 940 in the acylated segment of the RTX adenylate cyclase toxin (CyaA, ACT or AC-Hly) of Bordetella pertussis plays a key role in the interaction of the toxin with the target cell membrane. The aim of this study was to determine whether the corresponding conserved residues Y940, Y642, Y643 and Y639 secreted by the homologous RTX toxin CyaA from Bordetella bronchiseptica, HlyA from Escherichia coli, ApxIA from Actinobacillus pleuropneumoniae and RtxA from Kingella kingae play the same critical role in membrane insertion and pore formation. The hemolytic and cytotoxic activities of these toxins were completely impaired only after replacement of the conserved...
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Mechanism of action of bacterial toxins elevating the cAMP in host cells
Lepesheva, Anna ; Mašín, Jiří (advisor) ; Petráčková, Denisa (referee)
Cyclic adenosine monophosphate (cAMP) is an universal second messenger that regulates a large number of molecular mechanisms inside the eukaryotic cell. The level of synthesized cAMP is tightly regulated by endogenous adenylatecyclase (AC), and therefore this enzyme is often a target for various bacterial toxins. To manipulate intracellular cAMP levels in a target cell, bacteria have developed two different strategies for their toxins. Bordetella pertussis adenylate cyclase toxin (CyaA), Bacillus anthracis edema factor (EF) and Pseudomonas aeruginosa exotoxinY have in their structure an enzymatic AC domain which is activated by an intracellular cofactor and has several times higher activity than the eukaryotic AC enzyme itself. Other toxins, such as Bordetella pertussis pertussis toxin (PT), Vibrio cholerae cholera toxin (CT), and Escherichia coli heat labile toxin use ADP-ribosylation reaction of AC-coupled heterotrimeric G proteins to increase its activity and uncontrolled cAMP production. This work presents a literature search with accent on the molecular mechanism of interaction of these toxins with the target cell. Keywords: bacterial pathogens, virulence factors, intracellular cAMP elevation, bacterial toxins, adenylatecyclase (adenylylcyclase), Bordetella pertussis, Vibrio cholerae,...
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