Research predominantly on malaria parasites has been directed towards characterising, from a vaccine development viewpoint, surface antigens and their genes. Attempts at understanding the molecular biology of these amazing intracellular parasites has been much neglected. This thesis has focussed on understanding the molecular biology of two novel biological features in <i>Plasmodium falciparum</i>. Plasmodia rely on scavenging preformed host purines by the salvage pathway for nucleic acid synthesis. A pivotal enzyme in this pathway is hypoxanthine-guanine phosphoribosyltransferase (HGPRTase). The gene is present in a single copy in the <i>P.falciparum</i> genome and is located on a 1600 kb chromosome size molecule. The chromosomal copy of the HGPRTase gene has been cloned from the isolate K1 and also the cloned line HB3. DNA sequence analysis of the genes has been greatly hampered by their profound instability in conventional host-vector cloning and sequencing systems. However, some of these problems have now been surmounted and most of the gene sequenced. The picture that has emerged is fascinating. In all mammalian species examined, including man, the HGPRTase protein is encoded by less than 1kb of DNA. However, the genes span over a massive distance of 30-40 kb, the coding sequence is interrupted by eight intervening sequences which are present at precisely identical positions in all mammalian species. The organisation of the gene in <i>P.falciparum</i>, in contrast, is quite remarkable. It is contained within 2 kb of DNA and the coding sequence is interrupted by probably just a single intron at a position different from the introns in the mammalian gene. The coding sequence between different mammalian HGPRTase genes display over 90% sequence identity to each other, but only 70% identity to <i>P.falciparum</i>, most significantly the putative twenty residue catalytic domain of the protein is conserved. The monoclonal antibody, McAb 7.7, recognises a 34 kD parasite encoded protein exp-2 which has been characterised at the molecular level. The protein is synthesised as a 35 kD primary translation product which is presumably processed and exported outwith the parasite to reside in the parasitophorous vacuolar membrane and also vesicle-like structures in the erythrocyte cytoplasm. There is indirect evidence to suggest that vesicle-like structures are involved in trans-erythrocyte transport. The protein exp-2 has been affinity purified from differential detergent extracts. A number of proteins have been observed to reproducibly copurify with exp-2 of which one has been identified to be the erythrocyte anion-transporter band III. exp-2 has been convincingly demonstrated to be immunogenic in man. With a view to cloning the gene in expression libraries polyclonal antisera directed against exp-2 have been successfully raised. In conclusion the novel results that have emerged from this thesis are not only of great medical importance, but have widespread biological and evolutionary implications.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:653358 |
| Date | January 1990 |
| Creators | Khan, C. M. Anjam |
| Publisher | University of Edinburgh |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://hdl.handle.net/1842/11827 |
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