Hinds, L.; (2007) Identification and characterisation of a novel Plasmodium falciparum protein. Doctoral thesis , University of London.

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Abstract

The aim of this project was to identify and characterise a novel Plasmodium falciparum protein that plays a role in the invasion of host cells. Published genome, proteome and transcriptome data were used to select a novel protein for study. The data was filtered according to a set of criteria designed to identify uncharacterised proteins with features that would be expected of an invasion molecule, e.g. presence of a signal peptide and membrane anchor, as well as a late blood-stage expression profile. The gene product encoded by PF08_0008 was ultimately selected for study. Two regions of PF08_0008, from the N- and C-termini, were expressed in a recombinant form in E. coli and used to generate specific polyclonal antibodies in mice and rabbits. These antibodies have been used in a variety of experiments to characterise PF08_0008. Indirect immunofluorescence assays have shown that PF08_0008 is located at the parasite's apical end in both segmented schizonts and free merozoites. Immunoprecipitation and Western blot analyses have shown that PF08_0008 is proteolytically processed prior to schizont rupture. Fragments of PF08_0008 are detectable in culture supernatant, suggesting that shedding of this protein from the merozoite surface occurs. There is also preliminary evidence to suggest a role for PF08_0008 as an erythrocyte-binding molecule. The membrane anchorage of PF08_0008 has also been investigated, as it has sequence characteristics of both a type I membrane protein and a GPI-anchored protein. As part of this investigation, a line of parasites has been established in which endogenous PF08_0008 is expressed as a C-terminal fusion with a triple haemagglutinin epitope tag.

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