Support Core B: Proteomic Response to Toxic Perturbation
Dr. Dietmar Kültz, Associate Professor of Animal Science
Dr. Alan Buckpitt, Professor of Molecular Sciences
Dr. William Jewell, Facilities Manager for Advanced Instrumentation
The Core is developing and providing workflows for proteomics services including profiling of complex protein mixtures (with protein separation and mass spectrometric identification), analysis of posttranslational modification (PTM), including phosphorylation, thiol oxidation, ubiquitination, and activity-based protein profiling of proteases (caspases, etc.). In conjunction with the analytical core we also implement post-MS bioinformatic analyses to enable efficient and comprehensive processing, archival, and utilization of proteomic data sets. Experimental approaches practiced in this core are based on two-dimensional (2D) separation of complex protein mixtures by 2D gel electrophoresis (2DGE) and 2D liquid chromatography (2DLC). Fractionated mixtures are analysed by three different mass spectrometry approaches:
(i) analysis of trypic in gel digests from 2D gel spots and
(ii) analysis of complex peptide mixtures prepared by in-solution trypsin digestion by
(iia) offline LC-MALDI MS/MS or
(iib) online LC-MS/ MS. Proteomic datasets generated by these experimental approaches are then used for protein identification quantitative protein profiling, and PTM analysis.
In collaboration with the analytical core we also provide analytical support for storage and access of proteomics data. Additional integrating themes with other cores will attempt to find characteristic responses of human and other organisms to given agents, identifying biomarkers of early response, and finding no effect levels for the most sensitive biomarkers of effect. The emphasis is on increasing efficiency of resource utilization. Highly trained Core personnel concentrate their efforts on establishing innovative and robust workflows as well as performing technically demanding functions, providing help for planning experiments, and training Project personnel in conducting their own proteomics experiments where feasible.