This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor the Center, which is not necessarily the institution for the investigator.Technical Research Project 2, Protease Activity Imaging Technology (PAIT), will developtechnology for imaging of cellular proteolysis in three and four dimensions. Two independent andcomplementary approaches will be taken. First, a high throughput substrate phage display system willbe devised to identify highly selective indicator substrates for the vast majority of proteases encodedin the genome. Second, a chemical strategy will be applied to develop activity-based probes with anenzymatically activated fluorescent reporter. Both sets of beacons will be tested in biochemical andcellular models and will ultimately be used to image proteolysis at the cellular level
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