Combining Proteomic Technologies to Create a Platform for Spatiotemporal Enzyme Activity Profiling
EMSL Project ID
47293
Abstract
There is a need to investigate the functional activity of enzymes on the subcellular level to increase the understanding of a multitude of biological processes and protein interactions. The correlation of enzyme activity and location over time will provide valuable information about the function of subcellular compartments within the natural proteome. This project is developing spatiotemporal enzyme activity profiling (STEAP), which will combine activity-based protein profiling (ABPP) and cell fractionation for analysis by powerful and indepth LC-MS platforms already in place at the EMSL facility. The STEAP technology will provide a platform for assessing and comparing the functional activity of biological systems under various states and conditions.This project is directed toward the development of STEAP, and it will be applied initially to the human health relevant disease Invasive Aspergillosis (IA). Specifically, activity based probes (ABPs) will be synthesized to target glutamine synthetases, tyrosine phosphatases, serine hydrolases, cytochrome P450s, cysteine proteases, and glycoside hydrolases within the pathogenic Aspergillus fumigatus and an IA infection. ABPP of these systems will be combined with fractionation techniques to implement the STEAP platform. Analysis of samples by state of the art instrumentation at EMSL will provide high quality information about the functional activity of enzymes over space and time. This work will involve the development and demonstration of capabilities of STEAP, and reveal its utility to the study of systems biology.
Project Details
Start Date
2012-03-21
End Date
2014-09-30
Status
Closed
Released Data Link
Team
Principal Investigator
Co-Investigator(s)
Team Members
Related Publications
Sadler NC, MR Melnicki, MH Serres, ED Merkley, WB Chrisler, EA Hill, MF Romine, S Kim, EM Zink, S Datta, RD Smith, AS Beliaev, A Konopka, and AT Wright. 2013. "Live Cell Chemical Profiling of Temporal Redox Dynamics in a Photoautotrophic Cyanobacterium." ACS Chemical Biology 9 [Epub Ahead of Print](1):291-300. doi:10.1021/cb400769v
Wiedner SD, C Ansong, BJM Webb-Robertson, LM Pederson, S Fortuin, BA Hofstad, AK Shukla, EA Panisko, RD Smith, and AT Wright. 2013. "Disparate Proteome Responses of Pathogenic and Non-pathogenic Aspergilli to Human Serum Measured by Activity-Based Protein Profiling (ABPP)." Molecular & Cellular Proteomics. MCP 12(7):1791-1805. doi:10.1074/mcp.M112.026534. Epub 2013 Apr 18
Wiedner SD, KE Burnum, LM Pederson, LN Anderson, S Fortuin, LM Chauvigne-Hines, AK Shukla, C Ansong, EA Panisko, RD Smith, and AT Wright. 2012. "Multiplexed Activity-Based Protein Profiling of the Human Pathogen Aspergillus fumigatus Reveals Large Functional Changes Upon Exposure to Human Serum." Journal of Biological Chemistry.
Wiedner SD, LN Anderson, NC Sadler, WB Chrisler, VK Kodali, RD Smith, and AT Wright. 2014. "Organelle-Specific Activity-Based Protein Profiling in Living Cells." Angewandte Chemie International Edition 53(11):2919-2922. doi:10.1002/anie.201309135