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Title: Aspects of pyrimethamine resistance in the human malaria parasite Plasmodium falciparum
Author: Chan, Shiu-Wan
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 1991
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Pyrimethamine is a commonly used drug in prophylactic and therapeutic treatment of human malaria caused by the parasite Plasmodium falciparum. It has been postulated that the mode of action of pyrimethamine is by inhibition of the enzyme dihydrofolate reductase (DHFR). This was tested genetically in the first part of the study using a cross between a pyrimethamine sensitive and a pyrimethamine resistant cloned line 3D7 and HB3. The results of restriction-fragment-length-polymorphism (RFLP) analysis on the pattern of pyrimethamine resistance inheritance in the progeny confirm that the mutation causing resistance to pyrimethamine in HB3 is closely linked to the dihydrofolate reductase-thymidylate synthetase (DHFR-TS) gene and may be located within it, as previously inferred from sequencing and biochemical results. Structural alterations to DHRF resulting from point mutation in the DHFR gene has been suggested as one way in which resistance to pyrimethamine might occur. However, it is possible that other mechanisms e.g. over-production of the enzyme DHFR may also be important. The second part of the study investigated this possibility. Quantification of the enzyme level revealed no differences between the parents of the cross, 3D7 and HB3. However, pyrimethamine resistant mutant T9/94 (M1-1) selected in vitro was found to produce twice as much protein as the sensitive parent clone T9/94, suggesting that over-production of the enzyme can increase resistance to pyrimethamine. Over-productionof the protein was also observed in a naturally occurring pyrimethamine resistance clone PR70/CB3. Since a mutant over-producing the enzyme was identified, it was decicded to initiate studies that would ultimately be able to assess the role of transcriptional control in pyrimethamine resistance. As a first step, the 5' flanking regions of the DHFR genes from 3D7 and HB3 were sequenced and their transcriptional start sites mapped. Minor variations in the two sequences were found. The relationship of the sequence variations to the RFLP used in the genetic analysis is discussed.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available