Proteomic profiling of Nguni cattle liver tissue using gel and Gel-Free approaches: methodology development and potential applications

• Main Author: Buthelezi, Sindisiwe
• Other Authors: Blackburn, Jonathan
• Format: Dissertation
• Language: English
• Published: University of Cape Town 2014
• Subjects: Medical Biochemistry
• Online Access: http://hdl.handle.net/11427/3121

Includes abstract. === Includes bibliographical references. === In South Africa, resource-poor farmers mainly depend on livestock farming for their livelihoods, with cattle production being the most important livestock sector. As a consequence of natural selection in stressful conditions, Nguni cattle have been reported to be metabolically superior to other cattle breeds under unfavourable conditions. Using proteomics, with mass spectrometry at the core of the analysis, the objective of this study was to establish a reliable set of methods for the protein profiling of Nguni cattle livers. To achieve this several alternative technologies were employed and their outcomes compared namely, two-dimensional electrophoresis, fractionation by solution phase iso-electric focusing-reversed phase chromatography (IEF-RP), offline strong cation exchange- low pH reversed phase chromatography (SCX-RP) and offline high pH reverse phase-low pH reverse phase chromatography (RP-RP). All solution based methods were coupled to a tandem mass spectrometer. Protein identification was performed using the ParagonTMAlgorithm of Protein Pilot v4.0 as well as PEAKS v6. The IEF-RP and RP-RP methods achieved similar results in terms of number of proteins identified. In addition, proteins that play a role in the urea cycle (which is believed to contribute to the Nguni cattle’s enhanced metabolic ability) were all identified with both techniques. The RP-RP method was selected as the most appropriate method for future research linked to this work and will be used in the next phase of this project, on the basis that it is easier to automate compared to the IEF-RP method. It will be used beyond the scope of this work to compare levels of expression and modification of the liver proteins and their isoforms in Nguni and Hereford cattle grown under adverse environmental conditions, in order to identify those that may contribute to enhanced liver metabolism in Nguni cattle. This will be complemented by the identification and characterisation of potential polymorphisms with in such proteins that can be used to select for this trait during breeding.