| As instrumentation and mass spectrometry methods have advanced over the last century, the analyses of proteins and biological molecules of interest have evolved into the field of proteomics. Whole-cell analyses yielding thousands of protein identifications, coupled with relative quantification between multiple cell states, allows for a systems biology approach towards the identification of new drug targets, understanding biological systems and many other applications. In this body of work, a new mass spectrometry method for proteome profiling was used to compare protein expression between MCF-7 cells, a common breast cancer cell line, with and without tamoxifen treatment. The goal was to identify and characterize proteins which may be involved in the development of tamoxifen resistance. Another application of mass spectrometry in biological studies is the ability to pinpoint amino acid sites of covalent protein modifications. This in-turn lends further insight into many biological processes including signaling pathways, cytoskeletal polymerization, pathogenicity of bacteria and the reactivity of enzyme inactivating drug metabolites. This dissertation also explores the effect of ADP-ribosylation on peptide fragmentation patterns with both standard peptides and peptides modified by the bacterial toxin SpyA; ultimately identifying ions diagnostic of ADP-ribose and novel sites of modification on the cytoskeletal protein vimentin. Finally, mass spectrometry was used to identify a covalent adduct formed between the protease inhibitor ritonavir and P450 reductase following activation by cytochrome P450 3A4. |
