Proteomic analysis of protein complexes and cell cycle regulation in Trypanosoma brucei
type: CreativeWork
description: Trypanosoma brucei is a unicellular trypanosomatid protozoan parasite and the etiological agent of sleeping sickness in sub-Saharan Africa. The trypanosomatid order also includes the parasites Trypanosoma cruzi (Chagas disease) and Leishmania major (Leishmaniasis). Sleeping sickness is estimated to cause ~10,000 deaths per year and current treatments are expensive, difficult to administer and toxic. Although genomic sequencing of all three parasites has identified the coding sequences of these organisms, much is still unknown about protein function, with 64% of identified genes annotated as “hypothetical”, lacking obvious homology with proteins of known function. To further understand the unusual biology of this family of eukaryotes, this thesis aimed to provide evidence for protein function in Trypanosoma brucei in a high-throughput manner, utilising global proteomic analyses. This work has encompassed two main approaches: The global analysis of protein interactions and the analysis of proteome changes across the cell-cycle. To enable these approaches, I developed protocols for proteome wide analysis of protein complexes in Trypanosoma brucei, combining multiple forms of chromatography on ‘native’ lysates of cells to produce a proteome wide map of core, soluble protein complexes in this organism. I further performed preliminary studies to optimise in vivo formaldehyde crosslinking in T. brucei in order to characterise membrane bound protein complexes. I also developed methodologies to produce large populations of procyclic T. brucei cells highly enriched in different phases of the cell-cycle for proteomic analysis. In conjunction with the optimisation of methods for isobaric tag quantitation on Fusion mass spectrometers, I provide the first characterisation of protein regulation during cell division in T. brucei at an unparalleled proteomic depth. Together, these datasets provide a wealth of information about the interaction and cell cycle regulation of many thousa…
abstract: Trypanosoma brucei is a unicellular trypanosomatid protozoan parasite and the etiological agent of sleeping sickness in sub-Saharan Africa. The trypanosomatid order also includes the parasites Trypanosoma cruzi (Chagas disease) and Leishmania major (Leishmaniasis). Sleeping sickness is estimated to cause ~10,000 deaths per year and current treatments are expensive, difficult to administer and toxic. Although genomic sequencing of all three parasites has identified the coding sequences of these organisms, much is still unknown about protein function, with 64% of identified genes annotated as “hypothetical”, lacking obvious homology with proteins of known function. To further understand the unusual biology of this family of eukaryotes, this thesis aimed to provide evidence for protein function in Trypanosoma brucei in a high-throughput manner, utilising global proteomic analyses. This work has encompassed two main approaches: The global analysis of protein interactions and the analysis of proteome changes across the cell-cycle. To enable these approaches, I developed protocols for proteome wide analysis of protein complexes in Trypanosoma brucei, combining multiple forms of chromatography on ‘native’ lysates of cells to produce a proteome wide map of core, soluble protein complexes in this organism. I further performed preliminary studies to optimise in vivo formaldehyde crosslinking in T. brucei in order to characterise membrane bound protein complexes. I also developed methodologies to produce large populations of procyclic T. brucei cells highly enriched in different phases of the cell-cycle for proteomic analysis. In conjunction with the optimisation of methods for isobaric tag quantitation on Fusion mass spectrometers, I provide the first characterisation of protein regulation during cell division in T. brucei at an unparalleled proteomic depth. Together, these datasets provide a wealth of information about the interaction and cell cycle regulation of many thousa…
author: Crozier, Thomas William Monteiro
language: eng
keywords: Trypanosoma brucei, Proteomics, Cell cycle, Protein complexes
date published: 2016
date modified: 2026-07-24T02:08:19Z
provider: University of Dundee
publisher: Networked Digital Library of Theses and Dissertations
type: WebPage
source organization: University of Dundee