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From outside to inside : the relationship between eukaryote-like serine/threonine kinase signalling and antibiotic resistance in Enterococcal pathogens
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Sambrook, Megan (2022) From outside to inside : the relationship between eukaryote-like serine/threonine kinase signalling and antibiotic resistance in Enterococcal pathogens. PhD thesis, University of Warwick.
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Official URL: http://webcat.warwick.ac.uk/record=b3903358
Abstract
Enterococcus faecalis and Enterococcus faecium are opportunistic pathogens on the WHO priority pathogen list and are among the leading causes of nosocomial infections. Like many bacteria, they respond to changing environmental conditions using two-component systems (TCS) to initiate signal transduction pathways. Some of these networks mediate antibiotic resistance mechanisms, and the proteins involved are attractive drug targets. Additionally, the less abundant, eukaryotic-like serine-threonine kinases (eSTKs) are essential for sensing external stimuli and generating an intracellular response. Enterococci have a single eSTK called Irek, which has been previously implicated in the mediation of resistance to cephalosporins. Previously, the Roper lab has shown a direct linkage between Irek and the two TCS' that control vancomycin and intrinsic cephalosporin resistance. Eliminating Irek function via genetic deletion or chemical inhibition significantly increases vancomycin and cephalosporin susceptibility in multidrug resistant Enterococcal strains.
The research conducted in this thesis describes the study of these signalling pathways to identify novel drug targets. Various methods were tested to observe the noncanonical phosphorylation undertaken by TCS proteins to understand the relationship between vancomycin and cephalosporin resistance mechanisms. Additionally, the strong relationship between Irek and antibiotic resistance makes it an exciting drug target for antibiotic adjuvant therapeutics. An Irek activity assay was developed and optimised to identify novel Irek inhibitors. This assay was utilised at LifeArc to test over 7500 kinase inhibitor compounds in a high-throughput screen. One hit, AZD5438, was identified to inhibit Irek with micromolar potency. This compound was tested in an optimised bacterial growth assay alongside clinical concentrations of cefotaxime, to which E. faecalis is intrinsically resistant. AZD5438 successfully inhibited vancomycin-resistant E. faecalis growth as an antibiotic adjuvant. Further structural-activity relationship studies could improve bacterial specificity and turn this compound into a leading drug candidate.
Item Type: | Thesis (PhD) | ||||
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Subjects: | Q Science > QP Physiology Q Science > QR Microbiology R Medicine > RM Therapeutics. Pharmacology |
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Library of Congress Subject Headings (LCSH): | Drug resistance in microorganisms, Enterococcal infections, Cellular signal transduction, Drug targeting, Protein kinases, Phosphorylation | ||||
Official Date: | September 2022 | ||||
Dates: |
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Institution: | University of Warwick | ||||
Theses Department: | Warwick Medical School | ||||
Thesis Type: | PhD | ||||
Publication Status: | Unpublished | ||||
Supervisor(s)/Advisor: | Roper, David I. ; Kettleborough, Katy | ||||
Sponsors: | Medical Research Council (Great Britain) ; Warwick Medical School ; LifeArc | ||||
Format of File: | |||||
Extent: | 243 pages : illustrations (some colour), charts | ||||
Language: | eng |
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