| The use of mass spectrometry has been increasing in biological sciences over the last few decades as the technology has been getting more sensitive and specialised. From intact analysis of proteins to quantification of peptides or small molecules, the use of mass spectrometry continues to increase as it spreads through more disciplines in biological sciences. Here, four different questions were attempted to be answered starting with the less complex of identifying hydroxysterols in plasma as a method of identifying changes in the blood stream. Within the plasma ratios were created of 24S- and 27- hydroxycholesterol with cholesterol acids to create a relative quantification technique for analysis. The complexity increases with the identification of proteins involved in the synthesis of cholesterol and its metabolites in rat brain followed by identifying potential proteins involved in the synthesis of a UV absorbing molecule shinorine in the cyanobacteria Anabaena variabilis. Both proteomics analyses were able to identify some of the proteins involved in their respective pathways, while the cyanobacteria also quantified the levels of shinorine that were present by HPLC. While a few proteins were identified, it becomes obvious that more separation and sensitivity is needed to identify more of the pathways as they are not in high abundance in their respective proteomes. The final analysis performed was to understand the interaction of the protein STAT3beta with a known and published inhibitor, STATTIC. Using in-gel digest, STATTIC was found to bind to a cysteine residue away from the SH2 domain of STAT3beta. Within each question, mass spectrometry has been shown to be a valuable tool, however it is also shown to have drawbacks. It is shown most valuable when used to complement other techniques such as NMR, SDS-PAGE and Western blots, fluorescence assays and HPLC. |
