As information about the processes that regulate the spatial and temporal aspects of signal transduction through protein kinases becomes available, advanced techniques to analyze and apply these findings becomes necessary. The Proteomics/Beta ((3) Cell Core will work closely with the members of this PPG to provide proteomic analysis and characterization of PKA, PKC, AKAPs, as well as routine protein purification. Additionally, the core will prepare and analyze fetal and adult human islets which will be used by the investigators to apply their biochemical findings to a system relevant to the study of diabetes. The facility will provide excellent staff, resources, and state-of-the-art technologies to this PPG and work closely with each of the investigators to provide training in proteomics and primary islet cell cultue. Additionally, this Core will directly collaborate with the Imaging Core (National Center for Microscopy and Imaging Research (NCMIR) lead by Dr. Ellisman) and the Mitochondria Core (lead by Anne Murphy) to catalog the location of PKA and PKC signaling proteins in the mitochondria and to localize these signaling proteins in human islets. Specifically, the Proteomics/Beta (B) Cell Core will provide three essential services to the reaserchers: A) Proteomics Analysis: 1) Two dimesional electrophoresis/mass spectrometry for protein identification 2) Multidimensional Protein Identification Technology (MudPIT) 3) Protein expression profiling for fetal and human adult islets 3) Identification of protein:protein interactions 4) Analysis of post translational modifications B) Protein Purification and Expression 1) Routine large-scale purification of commonly used proteins 2) Specialized techniques for low expressing/difficult constructs 3) A centralized repository for all plasmids, proteins, and antibodies used by the investigators C) Preparation and Cultue of Humam Fetal and Adult Islets 1) Routine preparation, culture, and expansion of human fetal and adult islets
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