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Title: Identification and characterisation of secreted proteins of T. lestoquardi schizonts
Author: Chai, Z.-W.
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2005
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T. lestoquardi is an important tick-borne parasite that infects leukocytes of sheep and goats. The disease is mainly caused by the schizont stage which induces transformation of the infected cell to a state of uncontrolled proliferation. The purpose of this study was to identify schizont secreted proteins, which are likely to include molecules targeted by the immune response as well as those associated with the transformation process. An efficient method for the lysis and purification of schizonts from infected cells was established. This involved use of complement treatment to release schizonts from infected cells and an optimal density of Nycoprep gradient medium to separate schizonts from host cell components. An optimised method for metabolic labelling of schizont-infected cells was also established. Putative secreted proteins were detected from metabolically labelled schizont culture supernatants by SDS-PAGE and autoradiographic analysis. Using two-dimensional electrophoresis, mass spectrometry, including MALDI­TOF and Q-TOF, and N-terminal sequencing analysis, three putative secreted proteins of parasite origin were identified, namely Theileria mhsp70 (TLS3), hsp70 (TLS15) and inorganic pyrophosphatase (TLS16). Seven host-derived proteins were also identified, namely mhsp70 (TLSl), hsp60 (TLS2), mitochondrial ribosomal protein L12 (MRPL12, TLS4), ATP synthase-d (TLS5), ATP synthase 6(TLSll), PDH-Elb (TLS12) and A TP-dependent proteinase SP-22 (TLS13). Further investigation of the relationship of the host-derived proteins with the schizont using dual-labelling and confocal microscopy analysis provided evidence that the host-derived protein, MRPL12, is associated with intracellular schizonts with some co-localising with schizont mitochondria. The analysis also revealed that some host mitochondria are closely associated with the schizonts of T. lestoquardi.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available