MacCoss, Michael JMudge, Miranda Cristen2024-10-162024-10-162024Mudge_washington_0250E_27501.pdfhttps://hdl.handle.net/1773/52587Thesis (Ph.D.)--University of Washington, 2024Bacteria exist in every environment studied on Earth due to their wide-ranging metabolic capacities. In recent years, many studies have highlighted the significant contributions of bacteria as they interact with and respond to both other organisms and their environment through chemical signals. Because bacteria quickly respond to small physical and/or chemical environmental changes by adjusting their proteome, mass spectrometry-based proteomic techniques have gained momentum as a means to study the metabolic changes of mixed communities to environmental perturbations. In the following chapters, I utilize three different mass spectrometry techniques to identify the unique ways that microbes overcome environmental challenges in three unique projects. The complexity of the mass spectrometry data acquisition strategy utilized increases with chapter number, moving from the identification of the most abundant proteins in a single-organism culture to the eventual discovery of peptides from a complex ocean microbiome that can be isolated and targeted with mass spectrometry. In Chapter 1, I outline the many decisions that should be considered when conducting a complex proteomic or metaproteomic study. In Chapter 2, I describe adaptations of a model psychrophilic bacterium to subzero, hypersaline conditions to analyze the long-term effects of temperature, salinity, and nutrients on viability and survival and identify potential biomarkers for life detection through enrichment analyses of short amino acid chains. Chapter 3 transitions to a more complex biological system and quantitative mass spectrometry techniques. A high-resolution time series analysis of the bacterial metaproteome preceding a harmful algal bloom was utilized to isolate peptide biomarkers through quantitative discovery and targeted validation mass spectrometry analyses. In Chapter 4, the same microbiome was further characterized to elucidate metabolic patterns in the context of multiple algal blooms. Metabolic scheduling, the idea that bacteria prioritize distinct pathways at specific times of day, was explored to reveal functions that reflect global changes in the metabolic strategy of the microbiome between blooms. Finally, in Chapter 5 I reflect on the challenges still present in microbiome research, specifically in the context of metaproteomics, and discuss future directions for the research.application/pdfen-USCC BY-NDharmful algal bloommass spectrometrymicrobiomeocean bacteriaproteomicsBiological oceanographyMicrobiologyMolecular biologyMolecular and cellular biologyThesis