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Investigating GAP45 localisation and phosphorylation during plasmodium falciparum schizont development

Mohd Abd Razak, MRB; (2012) Investigating GAP45 localisation and phosphorylation during plasmodium falciparum schizont development. Doctoral thesis , UCL (University College London). Green open access

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Abstract

The invasion of erythrocytes by merozoites is driven by an actomyosin motor assembled below the parasite’s plasma membrane, with the myosin anchored on the inner membrane complex (IMC). The myosin (MyoA) is within a protein complex that is comprised of several proteins including myosin tail domain interacting protein (MTIP) and glideosome associated proteins (GAP) 45 and 50. A ternary complex of MyoA, MTIP and GAP45 is formed and later associates with GAP50. GAP45 is acylated by both myristoyl- and palmitoyl-fatty acids and is phosphorylated. This study has highlighted the GAP45 phosphorylation by calcium dependent protein kinase 1 (CDPK1) in vitro and its possible roles in schizogony. By site directed mutagenesis, substitution of S31, S89, S103 and S156 to alanine decreased the level of GAP45 phosphorylation, with S103A exhibiting a major decrease in 32P incorporation. Phosphorylation on S89 and S103 was studied further in parasites as both residues were among the phospho-sites in phosphopeptides identified in vivo. This study also showed that full length GAP45 labelled internally with GFP (FL-GAP45) is assembled into the motor complex, phosphorylated and transported to the developing IMC in early schizogony, where it accumulates during intracellular development until merozoite release. The C-terminal truncated GFP-GAP45 (N-GAP45; residues 1-29) localised at the plasma membrane instead of the IMC and was not assembled into the motor complex. The N-terminal truncated GFP-GAP45 (C-GAP45; residues 30-205) behaved like FL-GAP45. Modifying serine residues, S89 and S103, in GAP45 with alanine or aspartate had no apparent effect on its assembly into the protein complex or its intracellular location during schizont development and merozoite maturation. A second highly phosphorylated component of the complex (GAP40) was also identified. The early assembly of the motor complex suggests that it has functions in addition to its role in erythrocyte invasion.

Type: Thesis (Doctoral)
Title: Investigating GAP45 localisation and phosphorylation during plasmodium falciparum schizont development
Open access status: An open access version is available from UCL Discovery
Language: English
Additional information: Copyright restricted material has been removed from the digital copy of this e-thesis
UCL classification: UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences
URI: http://discovery.ucl.ac.uk/id/eprint/1354933
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