Date of Award
Doctor of Philosophy
MICHAEL D. BEST
SHANE FOISTER, BIN ZHAO, JEFFREY BECKER
Signaling lipids such as diacylglycerol (DAG) and the phosphatidylinositol polyphosphates (PIPns) play crucial roles in numerous cellular pathways. However, characterization of their activities is hindered by the complexity of associated signaling pathways and of the membrane environment. To address this issue, we have developed lipid probes that are effective for characterizing biological events using different applications, including activity-based probing (PIPns and DAG) and microarray analysis (PIPns). The activity-based probes have been applied to label receptor targets in multiple cancer cell proteomes through photocrosslinking followed by click reactions. The probes were found to label several proteins, as judged by on-gel fluorescence, and labeling was abrogated through various controls, such as heat denaturation and competition. Proteomic studies have been successfully performed to identify protein targets through biotin enrichment followed by mass spectrometric analysis. For microarray analysis, functionalized PIPn probes were synthesized and applied to develop a high throughput microarray analysis to measure protein-lipid binding affinity. These approaches will be invaluable for characterizing PIPn/DAG-regulated events and their involvement in disease. The design, synthesis and application of these lipid probes are included in this dissertation. In addition, the design and synthesis of other lipid probes are discussed, such as bis(monoacylglycero)phosphate (BMP), and lysophophatidylcholine (LPC) analogs.
Rowland, Meng Meng, "Chemical Tools to Characterize Membrane-Protein Binding Interactions Using Synthetic Lipid Probes. " PhD diss., University of Tennessee, 2011.