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Studies on the catalase genes and the acid tolerance response of mycobacteria.
thesisposted on 2015-11-19, 09:10 authored by Stephen. Gordon
Members of the genus Mycobacterium are major pathogens of man and animals. Tuberculosis causes almost 3 million deaths per year, currently making it the leading cause of death among bacterial infectious diseases. However, our knowledge of these pathogens is limited. This thesis investigated two questions (i) the role of mycobacterial catalases in isoniazid sensitivity, and (ii) the presence of an acid tolerance response. The mycobacterial M-catalase is thought to play a role in isoniazid sensitivity. Bacterial catalase enzymes are highly conserved at the amino acid level. An experimental strategy to clone the mycobacterial M- catalase gene sought to exploit this feature. Degenerate oligonucleotide primers were designed, based on previously cloned catalase genes, in an attempt to amplify the mycobacterial catalase gene using the polymerase chain reaction (PCR). Despite repeated attempts, this approach was unsuccessful. An attempt to identify the gene from a mycobacterial library using the catalase gene of Escherichia coli (katE) as a hybridisation probe also failed. Resistance to acidic environmental conditions is important for many mycobacterial species. Evidence was sought for the existence of an adaptive acid tolerance mechanism in the mycobacteria. To identify any genes that showed increased transcription in response to a decrease in extracellular pH, a promoter probe library was constructed with bacterial luciferase as the reporter. A clone (pSGA197) was identified that showed increased bioluminescence at the adaptive pH. The mycobacterial DNA insert in this clone was sequenced. Examination of the sequence revealed that the insert's putative acid-regulated promoter was located downstream from an open reading frame that coded for a possible sensor-kinase protein.
Date of award1995-01-01
Awarding institutionUniversity of Leicester