Research

Mass Spectrometry / Proteomics / Instrument Development

Our group focus is in biological mass spectrometry and proteomics research. Mass Spectrometry is currently changing the way that global or systems-level research on biological organisms is carried out. Mass spectrometry has enabled the field of proteomics to arise and provide a comprehensive view of all proteins present in a biological system under a given set of conditions. Our research centers on development and application of cutting-edge mass spectrometry for advanced proteomics research.

We are developing new methods and instrumentation that allow improved application of Fourier Transform Ion Cyclotron Resonance Mass Spectrometry (FTICR-MS) including custom ionization sources and transmission optics for increased sensitivity, new ion detection strategies that further improve mass resolution, accuracy and sensitivity. We are also developing new chemical strategies to target protein posttranslational modifications and protein-protein interactions. This combination of chemistry, biology and ion physics research is currently providing new information that enables a greater understanding in areas such as cystic fibrosis, bacterial drug resistance, cancer, and host-pathogen interactions.

Meeting Presentations

Bruce Lab presentations from National and International Meetings can be found on the Presentations page.

Further reading

More information on these topics and previous Bruce Lab research focuses can be found on our Publications page.

News

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Ongoing Research Projects

Quantitation of Protein Interactions in Cancer Cells

Protein interactions are key determinants of protein function in biological systems. Despite the potential that quantitative protein interaction information could have for all areas of cancer research, unbiased or large-scale quantitation of protein interactions within native living systems is a challenge that is unmet by today's technology. The capacity to identify and quantitate protein interactions on a large-scale within native cells, patient samples, or tissues does not currently exist. Improved capabilities to quantitate protein interactions will have a major impact on the understanding of cancer, metastasis and the development of anti-cancer drug resistance. This project aims to develop and apply quantitative cross-linking with cancer cells with advanced Protein Interaction Reporter (PIR) technology. Stable Isotope Label of Amino acids in Cell culture (SILAC) will be combined with PIR technology to allow quantitation of protein levels and protein interactions in cells. These capabilities will be applied to cisplatin-, taxol-, and SN-38 resistant cancer cells to allow quantitation of interactions relative to drug sensitive cancer cells. This project will provide the first relative quantitation data on protein interactions in cancer cells and the first unbiased measurements of functional regulation at the protein interaction level relevant to drug resistance.

Investigating Treatment Resistance Mechanisms in Chronic Bacterial Infections

Chronic bacterial infections are inherently resistant to treatment. This is true even if organisms are antibiotic-sensitive, and high concentrations of drugs reach infection sites. The infections that afflict patients with the genetic disease cystic fibrosis (CF) are a prime example. Once infection is established it cannot be eradicated, and lung dysfunction caused by chronic infections claim the lives of the vast majority patients. Importantly, the mechanisms producing treatment resistance in CF and other chronic infections are poorly understood. Here we exploit the infrastructure of an ongoing clinical trial, and new findings about genetic diversity within infecting P. aeruginosa populations as tools to study treatment resistance. In preliminary studies, we found that infecting Pseudomonas aeruginosa strains evolve to produce genetically diverse (but clonally-related) bacterial populations. We found that the relative abundance of subpopulations change as patients are treated with antibiotics. This is important because subpopulations that increase in abundance during treatment possess phenotypes that enable them to resist treatment in vivo, while subpopulations that decrease lack resistance functions. Studying these subpopulations could identify the mechanisms used by bacteria to withstand antibiotics in chronic human infections.

Protein Interactions of Genes of Unknown Function

Proteins are functional molecules in living systems and carry out most beneficial and deleterious function that affects life. A single protein may be involved in many different functional pathways and in living cells, interactions among proteins are the single strongest determinants that drive function. Therefore, the greatest opportunity to map coordinates of protein function for unknown or uncharacterized gene products would be presented if protein interaction networks that involve these proteins could visualized. The Protein Interaction Technology Core in this project is the product of many years of development efforts to pioneer new capabilities for visualization of protein interaction networks in live cells. This core will provide novel cross-linking and protein interaction identification technologies to support the overarching goal to place uncharacterized gene products within protein interaction networks in live A. baumannii cells. Through the core efforts to identify cross-linked peptides, interacting partner proteins and topological features of these interactions will be visualized. These data will form the basis of new interaction networks that can be mapped onto existing crystal structures for homologous or orthologous proteins and existing networks in other organisms to help link uncharacterized genes.

FT-ICR Instrumentation Development

The major challenges in proteomics research relate to the complexity of samples and the dynamic range over which measurements must be performed. In general terms, these demands are far greater than encountered in genomics since, protein abundances can vary more than genes, proteins have wider diversity in physical properties, and proteins have no means for signal amplification, nor analogous Watson-Crick base pairing as genes do. Thus the field of proteomics employs technology largely based on mass spectrometry measurements for large-scale protein identification and quantitation. However, since each protein can produce 50-100 peptides on average, measurements of peptide mixtures are extremely complex. In addition, current data-dependent measurement strategies lead to significant compression of achievable dynamic range, since such MS/MS measurements are only normally feasible on higher abundance peptides. Ideally, every peptide in a proteome-wide digest would be subjected to MS/MS to gain maximal information from proteomics experiments. This project will advance capabilities for large-scale proteomics through the development of a mass spectrometer array capable of MS/MS acquisition an order of magnitude faster than current state-of-the art technology. The MS array technology to be developed under this project will involve ion cyclotron resonance mass spectrometry that will yield higher mass resolving power, higher mass measurement accuracy as well as higher throughput acquisition. As a result, an order of magnitude or more peptides can be identified during a given experiment which will dramatically increase the information content of each proteomics analysis as well as the dynamic range of proteins that can be studied.

Publications

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Software

Our latest software can be downloaded from our Software page.