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Investigation of the protein interactions involved in the acylation of Escherichia coli α-hemolysin
Nemčeková, Lucia ; Osičková, Adriana (advisor) ; Šulc, Miroslav (referee)
Uropathogenic bacteria Escherichia coli are the primary cause of urinary tract infections. One of the virulence factors of these bacteria is α-hemolysin (HlyA), a protein belonging to the cytolytic RTX (Repeats in ToXin) toxins secreted by some gram-negative pathogenic bacteria. RTX toxins share several characteristic structural and functional domains and segments: (1) an N-terminal hydrophobic pore-forming domain, (2) an acylated segment where two lysine residues are modified by fatty acid chains, (3) a repetitive (RTX) domain binding calcium ions, and (4) a C-terminal secretion signal recognized by the type 1 secretion system. HlyA is synthesized as an inactive protoxin (proHlyA), which is activated by covalent acylation of the ε-amino groups of two conserved lysine residues, K564 and K690, by the co-expressed acyltransferase HlyC. Acyl-acyl carrier protein (acyl-ACP) serves as the acyl chain donor. However, the molecular mechanism by which the acyltransferase HlyC interacts with acyl-ACP and proHlyA is currently poorly understood. The aim of this bachelor thesis was to identify amino acid residues involved in the interaction between HlyC and ACP proteins. Based on an in silico interaction model of HlyC and ACP, positively charged residues in HlyC and negatively charged residues in ACP were...
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Investigation of the acyltransferase RtxC interaction with the Kingella kingae toxin RtxA
Lichvárová, Michaela ; Osičková, Adriana (advisor) ; Černá, Věra (referee)
The bacterium Kingella kingae was first isolated in 1960 by microbiologist Elizabeth O. King and until recently, it was considered a rare cause of human disease. However, over the past 30 years, an increasing number of papers have shown that this bacterium is an important paediatric pathogen, mainly affecting children aged 6 months to 3 years, causing mainly septic arthritis, osteomyelitis, infective endocarditis, and bacteraemia. K. kingae displays a strong cytotoxic effect against a variety of host cell types, which is caused by the secreted cytolysin RtxA, a member of the RTX (Repeats in ToXin) family. RtxA binds to glycosylated structures of the host cell, subsequently inserts into its cytoplasmic membrane, and forms cation-selective pores, leading to disruption of ion homeostasis and lysis of the attacked cell. RtxA is produced as an inactive protoxin proRtxA. Its activation is mediated by the acyltransferase RtxC, which transfers acyl chains to conserved lysine residues K558 and K689 in the protoxin. It uses the acyl carrier protein ACP as the acyl donor. Currently, it is unclear how the RtxC, acyl-ACP, and proRtxA proteins interact with each other and which amino acid residues are responsible for these interactions. The aim of this thesis was to identify the residues responsible for these...
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