Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.651478
Title: Target protein mutation in the analysis of bactericidal antibiotic action and resistance
Author: Gill, Anne Elizabeth
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2002
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Abstract:
Future antimicrobial drug design and resistance minimisation, requires an understanding of current antimicrobial action and resistance. This study investigated bactericidal action and resistance within two antimicrobial groups: the aminoglycosides and the fluoroquinolones. The requirement of protein synthesis in bactericidal aminoglycoside action was initially analysed. Pre-incubation with chloramphenicol significantly reduced streptomycin bactericidal activity, indicating a role of protein synthesis in cell death. Substituting streptomycin (1mg/L) for chloramphenicol gave a similar significant reduction in bactericidal activity, indicating that at sub-lethal concentrations streptomycin acts as a protein synthesis inhibitor which blocks its own cell death when challenged with a higher concentration. Streptomycin induced cell death thereby requires additional protein synthesis dependent death mechanisms. Stationary phase cells were resistant to drug challenge, supporting this result. The role of Ribosomal S12 mutations to aminoglycoside action and resistance was characterised in a step-wise mutation study. The S12 mutation Lys87®Arg confers streptomycin resistance of >1500mg/L in a single step selection; however the role of this S12 in resistance to other aminoglycosides has not been previously characterised. Mutants of E. coli NCTC10418 were selected in four steps (I-IV) on aminoglycoside containing plates. Lack of cross resistance between streptomycin mutants and other aminoglycosides (gentamicin, neomycin, kanamycin, tobramycin), suggested that the streptomycin target may not be shared by other aminoglycosides. Cross-resistance between neomycin, tobramycin and kanamycin mutants, implied that these drugs may share similar targets and binding pockets. PCR amplification and sequencing of the rpsL gene encoding the S12 protein demonstrated a Lys87®Glu mutation in streptomycin resistant mutants. Ribosomal S12 mutations were absent in gentamicin, neomycin, kanamycin and tobramycin resistant mutants. Replacement of Lys87 with a glutamic acid residue resulted in a streptomycin MIC of 25mg/L.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.651478  DOI: Not available
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