Strategies for Protein and Peptide Characterization and Quantification using Electron-Transfer Dissociation Mass Spectrometry and Intrinsic Fluorescence

Under the supervision of Associate Professor Joshua J. Coon At the University of Wisconsin-Madison The following chapters detail strategies for peptide and protein sequence analysis featuring electron-transfer dissociation (ETD) tandem mass spectrometry (MS/MS) and quantification using ultraviolet light-induced intrinsic fluorescence (UV-IF). Chapter 1 provides a brief background and overview. Chapter 2 discusses the optimization of the ETD MS/MS duty cycle for large-scale shotgun experiments. In Chapter 3, the first comprehensive analysis of peptide anions using negative ETD (NETD) is detailed. I report in Chapter 4 on the performance of an ion-ion reaction cell for intact protein analysis using large precursor populations for ETD MS/MS. The application of UV-IF for peptide detection and quantification using a custom fluorescence excitation and detection device is discussed in Chapter 5. An analysis of intact proteins from the 26S proteasome of Arabidopsis using top-down mass spectrometry and quantification by UV-IF is described in Chapter 6. In coordination with the Wisconsin Initiative for Science Literacy (WISL), Chapter 7 is a general description of my graduate research intended for non-specialists in an effort to promote science literacy in the broader community.