Characterisation of the plasma membrane subproteome of bloodstream form Trypanosoma brucei |
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Authors: | Bridges Daniel J Pitt Andrew R Hanrahan Orla Brennan Kiva Voorheis H Paul Herzyk Pawel de Koning Harry P Burchmore Richard J S |
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Institution: | Division of Infection and Immunity, Institute of Biomedical and Life Sciences, University of Glasgow, Glasgow, UK. |
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Abstract: | Proteome analysis by conventional approaches is biased against hydrophobic membrane proteins, many of which are also of low abundance. We have isolated plasma membrane sheets from bloodstream forms of Trypanosoma brucei by subcellular fractionation, and then applied a battery of complementary protein separation and identification techniques to identify a large number of proteins in this fraction. The results of these analyses have been combined to generate a subproteome for the pellicular plasma membrane of bloodstream forms of T. brucei as well as a separate subproteome for the pellicular cytoskeleton. In parallel, we have used in silico approaches to predict the relative abundance of proteins potentially expressed by bloodstream form trypanosomes, and to identify likely polytopic membrane proteins, providing quality control for the experimentally defined plasma membrane subproteome. We show that the application of multiple high-resolution proteomic techniques to an enriched organelle fraction is a valuable approach for the characterisation of relatively intractable membrane proteomes. We present here the most complete analysis of a protozoan plasma membrane proteome to date and show the presence of a large number of integral membrane proteins, including 11 nucleoside/nucleobase transporters, 15 ion pumps and channels and a large number of adenylate cyclases hitherto listed as putative proteins. |
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Keywords: | Bioinformatics Mass spectrometry Plasma membrane Protein separation Trypanosoma brucei |
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